Cell culture

Sequence-independent single-primer-amplification (SISPA) as a screening technique for detecting unexpected RNA viral adventitious agents in cell cultures

Published on: 12th March, 2021

OCLC Number/Unique Identifier: 8982622827

The sequence-independent, single-primer amplification (SISPA) enables the random amplification of nucleic acids, allowing the detection and genome sequencing of different viral agents. This feature of SISPA method provides evidence for application of it in monitoring the presence of adventitious RNA viruses in cell cultures. We evaluated SISPA method for the detection of a challenge RNA virus representing adventitious agent in cell cultures. Besides, by optimizing the SISPA method in our laboratory, we found false-positive results on negative control lanes in electrophoresis gels. To investigate the sources of contamination, false-positive results of SISPA were cloned into Escherichia coli cells, sequenced, and phylogenetically analyzed. This data revealed that the SISPA method can be used as an adjunct method to confirm the absence of unexpected adventitious RNA viruses in cell cultures. The phylogenetic analysis of SISPA contaminant sequences showed that the false-positive results were caused by nucleic acid amplification of commercial cDNA synthesis kit reagents, probably tracing back to expression plasmids and host ribosomal sequences, used for the production of enzymes. Therefore, laboratories using random amplification methods must be constantly aware of the potentials of such contaminations, yielding false-positive results and background noise in the final NGS reads.
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In vitro beneficial effects of a flax extract on papillary fibroblasts define it as an anti-aging candidate

Published on: 5th May, 2021

OCLC Number/Unique Identifier: 9045687547

Objective: During aging, skin undergoes structural, cellular and molecular changes, which not only alter skin mechanical properties but also biological and physiological functions. Structurally the epidermis becomes thinner, the dermal epidermal junction flattens and the extra-cellular matrix component of the dermis is disorganized and degraded. The dermis is composed of two compartments: The Reticular dermis is the deepest and thickest part while the upper layer, the papillary dermis, which is much thinner and is in close contact with epidermis, plays an important role in the structure and function of the skin. We have recently shown that the papillary dermis was preferentially affected by skin aging because the activity of fibroblasts in this region was especially altered as a function of age. The purpose of this study was to investigate the capacity of a flax extract as anti-aging component. Method: We investigated the capacity of a flax extract to stimulate or restore the activity of papillary fibroblasts from young and old donors in cultured monolayers and in reconstructed skin. Several biological markers of extracellular matrix homeostasis and mechanical properties were investigated. Results: The tested flax extract seemed to improve parameters known to change with age: I/ In monolayers after treatment the number of aged fibroblasts increased II/ In reconstructed skin the flax extract appears to positively regulate some biological activities; particularly in aged fibroblasts where the deposition of laminin 5, fibrillin 1, procollagen I were increased in the dermis and the secretion of specific soluble factors like MMP1, MMP3 and KGF were regulated to levels similar to those observed in young fibroblasts III/ Mechanical properties were improved particularly for elastics parameters (R5, R2 and R7). Conclusion: The flax extract is a promising anti-aging compound. The treatment of aged papillary fibroblasts resulted in a return to a younger-like profile for some of the studied parameters.
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Therapeutic application of herbal essential oil and its bioactive compounds as complementary and alternative medicine in cardiovascular-associated diseases

Published on: 10th March, 2020

OCLC Number/Unique Identifier: 8586050480

Background: Herbal essential oil contains pharmacological benefits for intervention treatment of various diseases. Studies have demonstrated its antimicrobial, antioxidant, and anti-inflammatory effect involving in vitro cell culture and preclinical animal models. It has been also traditionally used to reduce anxiety and hypertension in human. However, scientific studies elucidating its mechanism of action and pharmacological targets, as well as its effectiveness and safety as phytotherapeutic compounds are still progressing. Recent studies showed its promising effect in depression-cardiovascular disease intervention. However, comprehensive evaluations to enlighten latest advancement and potential of herbal essential oil are still lacking. Objective: In this systematic review, the depression-cardiovascular effects of herbal essential oil on lipid profile, biochemical and physiological parameters (e.g haemodynamic) are presented. The route of delivery and mechanism of action as well as main bioactive compounds present in respective essential oil are discussed. Methods: Article searches are made using NCBI PubMed, PubMed Health, SCOPUS, Wiley Online, tandfonline, ScienceDirect and Espacenet for relevant studies and intellectual properties related to essential oil, depression and cardiovascular disease. Results: In experimentation involving in vitro, in vivo and clinical trials, herbal essential oil showed its effectiveness in reducing coronary artery disease (narrowing of the arteries), heart attack, abnormal heart rhythms, or arrhythmias, heart failure, heart valve disease, congenital heart disease, heart muscle disease (cardiomyopathy), pericardial disease, aorta disease, Marfan syndrome and vascular (blood vessel) disease. Conclusion: This review gives a valuable insight on the potential of essential oil in the intervention of depression associated with cardiovascular diseases. Studies showed that herbal essential oil could act as vasodepressor, calcium channel blocker, antihyperlipidemia, anticoagulant, antiatherogenesis and antithrombotic. It can be proposed as an interventional therapy for depression-cardiovascular disease to reduce doses and long-term side-effect of current pharmacological approach.
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Protective functions of AEURA in Cell Based Model of Stroke and Alzheimer disease

Published on: 6th June, 2017

OCLC Number/Unique Identifier: 7317651488

Stroke and neurodegenerative diseases including Alzheimer’s disease (AD) are responsible for a major proportion of mortalities in the elderly. We have previously investigated novel mechanism-based therapies of AEURA in cell culture models against viral infection and in glutamate excitotoxity. In our new studies, we propose that the homeopathic formula AEURA could serve as a potential therapeutic agent for stroke & for AD. In examining AEURA treatment of PC12 cells exposed to glutamate excitotoxicity, hypoxia /re-oxygenation injury and A-Beta toxicity. We demonstrated an increased survival rate in AEURA treated cells by comparison to control cells. In examining the therapeutic potential of AEURA in PC12 cells this homeopathic agent was found to be neuroprotective against either glutamate induced toxicity, hypoxia /re-oxygenation stress or cell stress resulting from viral infection (with either HSV-1 or rhinovirus). Our ongoing studies involve examining the neuroprotective potential AEURA in vivo using rodent models of stroke & AD.
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