imagine (Institute of molecular genetics and genetic engineering)
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Doxorubicin and quercetin combined effect on SAOS-2 cells grown in 2D and 3D model systems
Get PDFOsteosarcoma (OS) is a highly aggressive primary malignant bone tumor that most commonly affects children, adolescents, and young adults. The standard treatment for OS consists of surgical resection and chemotherapy, whereas radiation therapy is recommended for the unresectable tumor. Due to its easy metastasis and recurrence, the 5-year overall survival rate is only 66.5 %. Thus, there is a critical need to recognize the molecular mechanisms underlying OS development and pathogenesis. Traditionally, two-dimensional (2D) cells are widely used in cancer biology and pre-clinical studies. However, 2D models are unable to mimic cell–cell and cell-extracellular matrix interactions which are crucial for adequate cellular function. Three-dimensional (3D) model systems are able to recapitulate key features of human cancer and are recognized as a promising platform for fundamental and translational research. In the present work, we established an osteosarcoma 3D model based on alginate microbeads and studied the effect of combined treatment with doxorubicin (Doxo), widely used chemotherapeutic, and quercetin (Quer), a plant pigment with anticancer properties, on OS model systems
Silver(I) complexes with voriconazole as promising anti-Candida agents
Get PDFRecognizing that metal ions play an important role in modifying the pharmacological properties of known organic-based drugs, the present manuscript addresses the complexation of the antifungal agent voriconazole (vcz) with the biologically relevant silver(I) ion as a strategy for the development of new antimycotics. The synthesized silver(I) complexes with vcz were characterized by mass spectrometry, IR, UV–Vis and NMR spectroscopy and single-crystal X-ray diffraction analysis. The crystallographic results showed that complexes {[Ag(vcz)(H2O)]CH3SO3}n (1), {[Ag(vcz)2]BF4}n (2) and {[Ag(vcz)2]PF6}n (3) have polymeric structures in the solid state, in which silver(I) ions have a distorted tetrahedral geometry. On the other hand, DFT calculations revealed that the investigated silver(I) complexes 1–3 in DMSO exist as linear [Ag(vcz-N2)(vcz-N19)]+ (1a), [Ag(vcz-N2)(vcz-N4)]+ (2a) and [Ag(vcz-N4)2]+ (3a) species, respectively. The evaluated complexes showed an enhanced anti-Candida activity compared to the parent drug with minimal inhibitory concentration (MIC) values in the range of 0.02–1.05 μM. In comparison with vcz, the corresponding silver(I) complexes showed better activity in prevention hyphae and biofilm formation of C. albicans, indicating that they could be considered as promising agents against Candida that significantly inhibit its virulence. Also, these complexes are much better inhibitors of ergosterol synthesis in the cell membrane of C. albicans at the concentration of 0.5 × MIC. This is also confirmed by a molecular docking, which revealed that complexes 1a – 3a showed better inhibitory activity than vcz against the sterol 14α-demethylase enzyme cytochrome P450 (CYP51B), which plays a crucial role in the formation of ergosterol
Novel formulation for maintaining the integrity of lipid vesicles (Peptosome)
Get PDFModern skin care formulations contain liposomes – nano-spherical vesicles. Liposomes are the best
delivery system for the controlled release of active and sensitive ingredients (e.g. vitamin C) into the
deeper skin layers. Formulations with liposomes have been shown to release vitamin C ten times faster
into the deeper skin layers, while protecting the vitamin C from oxidation and degradation. However, as
soon as the packaging of such a formulation is opened, its contents are exposed to atmospheric oxygen
and the ingredients, including the liposomes, oxidise and dry out. Our solution: PEPTOSOME extends the
shelf-life of expensive liposome-based skin formulations by stabilising both, the liposomes and their
contents.
The inspiration for PEPTOSOME lies in the ancient and remarkable phenomenon of resurrection plants.
Part of this dehydration survival strategy based on specific, flexible proteins has been implemented in
PEPTOSOME. As our invention PEPTOSOME is a formulation based on a flexible peptide structure, it
circumvents the lack of selectivity of small molecules and provides better affinity and coverage. The safety
and skin-beneficial function of the extracts of these proteins/peptides is proven. PEPTOSOME is
completely innovative, cost-effective, easy to transport (at room temperature), eco-friendly, and contains
no toxic chemicals. Our invention is also suitable for stabilising liposomes loaded with other skin-beneficial
substances (anti-ageing agents, moisturisers, etc.), suitable for personalised care. Finally, LEAPOSOME can
also be used with liposomes in the food industry, pharmacy, medicine and organic farming.Principal Investigator: Dr Marija Vidović, IMGGEDuration period: 2024-202
THE ROLE OF SOX2 AND SOX9 TRANSCRIPTION FACTORS IN THE REACTIVATION-RELATED FUNCTIONAL PROPERTIES OF NT2/D1- DERIVED ASTROCYTES
No full textAstrocytes are the main homeostatic cells in the central nervous system. During reactive
astrogliosis, these cells transform from quiescent into a reactive state by reacquiring some of
the precursor properties. This process, activated in response to different pathological conditions
of the brain, serves as a compensatory response that modulates tissue damage and recovery.
SOX2 and SOX9 transcription factors have important roles during gliogenesis. These
transcription factors are down-regulated in mature astrocytes and re-expressed in reactive
astrocytes, however, their roles in these cells are still not fully elucidated. We focused our study
on reactivation-related functional properties of astrocytes mediated by these proteins. Initial
screening of SOX2 and SOX9 expression after sensorimotor cortex ablation injury in rats was
performed using immunohistochemistry. Further gain-of-function studies were done in vitro
using astrocytes derived from the NT2/D1 cell line (NT2/A). Our results revealed the direct
involvement of SOX2 in the reacquisition of proliferation in mature NT2/A, while SOX9
overexpression increased migratory potential and glutamate uptake in these cells. Our results
imply that modulation of SOX gene expression may change the functional properties of
astrocytes, which holds promise for the discovery of potential therapeutic targets in the
development of novel strategies for neural tissue regeneration and recovery.VII Congress of the Serbian Genetic Society Zlatibor; October 2 to 5, 2024
Alginate microfibers with incorporated hydroxyapatite particles for 3D osteosarcoma cell cultures and rapid anticancer drug screening
No full textOsteosarcoma is the most common primary malignant bone tumor and requires effective
preclinical models to facilitate the development of new anticancer drugs. Current models
include two-dimensional (2D) cell cultures and animal models, which cannot often fully
replicate human cancer biology, leading to discrepancies in drug efficacy results between
preclinical studies and clinical trials. This work aimed to develop a 3D osteosarcoma model
based on alginate hydrogel with incorporated hydroxyapatite (HAP) particles, to create a
simplified, yet reliable system for rapid anticancer drug screening.
Osteosarcoma cells (murine cell line K7M2-wt) were suspended (4×106 cells cm-3
) in 2 wt. %
alginate solution with or without 2 wt. % commercial HAP powder. The suspensions were
manually extruded through a 26-gauge needle into a 0.18 M Ca2+ gelling solution. The obtained
microfibers were washed and cultured for up to 21 days. For anticancer drug testing, 3 cm of
microfibers per well in a 96-well plate were treated with 0.25-20 µM doxorubicin solutions,
while treated cells in monolayers served as controls. Histology (H&E staining) was used for the
analysis of cell morphology and distribution, while the cell metabolic activity and the halfmaximal inhibitory concentration (IC50) were determined by the MTT assay.
The 3D cultivation of osteosarcoma cells in alginate microfibers with and without HAP has
shown that the microfibers supported cell viability, metabolic activity, and formation of cellular
aggregates. The results of anticancer drug testing indicated higher resistance of the 3D cultures
to doxorubicin compared to that in 2D: the IC50 value for both 3D cultures of K7M2-wt cells
was 3 µM, while that for the 2D culture was 0.5 µM. These results are consistent with the
higher resistance to anticancer drugs observed in clinics compared to existing preclinical
models and indicate the importance of developing more reliable in vitro models to improve
preclinical platforms for drug discoveryTwenty-fifth Jubilee Annual Conference YUCOMAT 2024 & Thirteenth World Round Table Conference on Sintering XIII WRTCS 2024, Herceg Novi, Montenegro, September 2 to 6, 202
The Microbiome–Genetics Axis in Autism Spectrum Disorders: A Probiotic Perspective
No full textAutism spectrum disorder (commonly known as autism) is a complex and prevalent neurodevelopmental condition characterized by challenges in social behavior, restricted interests, and repetitive behaviors. It is projected that the annual cost of autism spectrum disorder in the US will reach USD 461 billion by 2025. However, despite being a major public health problem, effective treatment for the underlying symptoms remains elusive. As numerous literature data indicate the role of gut microbiota in autism prognosis, particularly in terms of alleviating gastrointestinal (GI) symptoms, high hopes have been placed on probiotics for autism treatment. Approximately twenty clinical studies have been conducted using single or mixed probiotic cultures. However, unequivocal results on the effect of probiotics on people with autism have not been obtained. The small sample sizes, differences in age of participants, choice of probiotics, dose and duration of treatment, outcome measures, and analytical methods used are largely inconsistent, making it challenging to draw distinctive conclusions. Here, we discuss the experimental evidence for specific gut bacteria and their metabolites and how they affect autism in light of the phenotypic and etiological complexity and heterogeneity. We propose a personalized medicine approach for using probiotics to increase the quality of life of individuals with autism by selecting specific probiotics to improve particular features of the condition
Greener extraction of intracellular poly(3-hydroxybutyrate) from Zobellella denitrificans biomass using enzyme cocktails and deep eutectic solvents
No full textThe accumulation of petroleum-based plastics, which degrade poorly,
highlights the need for sustainable alternatives. Biopolymers like
polyhydroxyalkanoates (PHAs), biodegradable and non-toxic polyesters
derived from renewable resources, offer a greener solution by
reducing plastic pollution and petroleum reliance. Among PHAs,
poly(3-hydroxybutyrate) (PHB) is the most studied and produced
biopolymer. It is accumulated within microbial cells and requires
efficient extraction and purification. Organic solvents like chloroform
and acetone are still golden standard for their downstream processing.
In our study, the bacterium Zobellella denitrificans MW1, engineered
to use waste glycerol, was used for PHB-rich biomass (24% w/w PHB)
production. We assessed the extraction of PHB using mixtures of
enzymes and natural deep eutectic solvents (NADES)
Ultra-Early Diffuse Lung Disease in an Infant with Pathogenic Variant in Telomerase Reverse Transcriptase (TERT) Gene
No full textThe pathogenic variants in the telomerase reverse
transcriptase (TERT) gene have been identified in adults
with idiopathic pulmonary fibrosis, while their connection to childhood diffuse lung disease has not yet been
described. Within this study, we present a case of a fivemonth-old, previously healthy infant, with early-onset
respiratory failure. The clinical suspicion of diffuse lung
disease triggered by cytomegalovirus (CMV) pneumonitis was based on clinical and radiological presentation.
Multiorgan involvement was not confirmed. Considering
the possible connection between CMV pneumonitis and
early-onset respiratory failure, clinical exome sequencing
was performed and a novel variant, classified as likely
pathogenic in the TERT gene (c.280A>T, p.Lys94Ter)
was detected. After segregation analysis yielded negative
results, the de novo status of the variant was confirmed.
Respiratory support, antiviral and anti-inflammatory therapy offered modest benefits, nevertheless, eighteen months
after the initial presentation of disease, an unfavourable
outcome occurred. In conclusion, severe viral pneumonia has the potential to induce extremely rare early-onset
diffuse lung disease accompanied by chronic respiratory insufficiency. This is linked to pathogenic variants in the
TERT gene. Our comprehensive presentation of the patient
contributes to valuable insights into the intricate interplay
of genetic factors, clinical presentations, and therapeutic
outcomes in cases of early-onset respiratory failure
Comparison of the effects of combined doxorubicin and quercetin treatment on 2D and 3D osteosarcoma model systems
No full textOsteosarcoma (OS) is a highly aggressive bone tumor primarily affecting pediatric patients,
with a 5-year survival rate of 66.5%. Standard treatments include surgical resection,
chemotherapy, and radiation for tumors that cannot be surgically removed. Literature data has
shown that quercetin, a plant pigment with known anti-tumor effects, could enhance
chemotherapeutic effect of doxorubicin on cancer cells. In this study we have investigated the
effects of the combined treatment with doxorubicin, a commonly used chemotherapeutic, and
quercetin, on human osteosarcoma cell line, SAOS-2 cultured in 2D and 3D model systems.
SAOS-2 (8 x 106 cells/ml) immobilized in alginate microbeads (~ 1 mm in diameter) showed
enhanced viability for up to 21 days after immobilization. In addition, cells were treated with
1 μg/ml Doxo and 5 μM Quer, as well as with each compound separately. Following treatment,
we assessed cell viability, morphology, and the expression of genes. Our results have shown
that the combined treatment statistically significantly decreased the viability of SAOS-2 cells
cultured in 2D and 3D conditions compared to cells treated with doxorubicin. We analyzed the
expression of genes associated with poor prognosis in patients such as pluripotency genes, an
OS marker, and a resistance-related gene. Collectively our results show different responses to
the combined treatment depending on the model system used. The combined treatment
substantially reduced cell viability in 2D and 3D models and decreased expression of genes
associated with poor prognosis compared to doxorubicin alone in 3D models. We can
hypothesize that microenvironment-based mechanisms modulate cell sensitivity to therapy and
increase resistance to treatment of osteosarcoma cells cultured in 3D condition. Further
research is needed to investigate quercetin’s possible role in tumor treatment.Twenty-Second Young Researchers Conference – Materials Science and Engineering
December 4 – 6, 2024, Belgrade, Serbi
Doxorubicin application in a 3D osteosarcoma cell culture model
No full textThe most common primary malignant bone tumor that predominantly affects children and
adolescents is osteosarcoma. Comprehensive osteosarcoma treatment involves neoadjuvant
multi-agent chemotherapy followed by surgical resection and adjuvant multi-agent
chemotherapy. Unfortunately, this complex treatment is insufficiently effective, as the tumor
recurs in over 40 % of cases. The development of new drugs for osteosarcoma is a complex
and slow process due to various limitations of preclinical drug evaluation methods. One
potential solution is to develop three-dimensional (3D) systems that replicate key in vivo
physiological conditions, using biomaterials as extracellular matrices and biomimetic
bioreactors to ensure efficient mass transport and adequate biophysical signals. This study
aimed to evaluate a previously developed 3D culture model for osteosarcoma cells based on
macroporous composite scaffolds (2 wt.% alginate and 2 wt.% hydroxyapatite) in conjunction
with a perfusion bioreactor for anticancer drug screening. Murine osteosarcoma cells (K7M2-
wt) were seeded onto the scaffolds (15x106 cells cm-3 of scaffold volume) and cultivated under
static conditions for one day. Two experimental studies were performed using the anticancer
drug doxorubicin. In the first study, following the static cultivation, doxorubicin (1 μg cm-3 of
culture medium) was applied for 1 day in a “3D Perfuse” bioreactor (Innovation Center of
Faculty of Technology and Metallurgy, Belgrade, Serbia) at a flow rate of 0.27 cm³ min-1,
corresponding to the superficial velocity of 40 μm s-1. In the second study, after the 1-day static
cultivation, cell seeded scaffolds were cultivated for 7 days under the same continuous flow
conditions, which allowed spontaneous formation of spheroid-like structures. Next,
doxorubicin was applied in the same manner as in the first study. The cultivated scaffolds in
both studies were then assessed regarding the cell metabolic activity using the MTT (3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The obtained results indicated
lower sensitivity of cells cultured in the 3D environment to the tested drug as compared to
those in 2D in agreement with chemotherapy resistance observed in patients.Twenty-Second Young Researchers Conference – Materials Science and Engineering December 4 – 6, 2024, Belgrade, Serbi