Recent advances on gradient hydrogels in biomimetic cartilage tissue engineering [version 2; referees: 2 approved]

Articular cartilage (AC) is a seemingly simple tissue that has only one type of constituting cell and no blood vessels and nerves. In the early days of tissue engineering, cartilage appeared to be an easy and promising target for reconstruction and this was especially motivating because of widespread AC pathologies such as osteoarthritis and frequent sports-induced injuries. However, AC has proven to be anything but simple. Recreating the varying properties of its zonal structure is a challenge that has not yet been fully answered. This caused the shift in tissue engineering strategies toward bioinspired or biomimetic approaches that attempt to mimic and simulate as much as possible the structure and function of the native tissues. Hydrogels, particularly gradient hydrogels, have shown great potential as components of the biomimetic engineering of the cartilaginous tissue

Editorial: Microenvironment-Derived Stem Cell Plasticity

No abstract available

Stimulating Innovations from University through the Use of Digital Fabrication - Case Study of the SciFabLab at Faculty of Mechanical Engineering, University of Belgrade

Fabrication laboratory (FabLab), first created at MIT-USA, is a digital fabrication workshop equipped with tools that enable students to produce their own prototypes and engage in “learning-by-doing” process. Fablab is a do-it-yourself "DIY" workshop with open-source digital design and manufacturing CNC-machines (3D printers, laser cutters and electronics e.g. Arduino, Raspberry Pi), with a strong focus towards technology transfer options and as such is strongly connected with both the small and medium entreprizes (SMEs) and large industry in a unique ecosystem [1]. There are no such ecosystems yet in Serbia. A project started at the University of Belgrade - Faculty of Mechanical Engineering, already possessing fablab-ready infrastructure [CNC (computer-numerical controlled) machines, lab/workshop space and teaching venue] aims to establish one component of such an ecosystem –a Scientific Fab Lab (SciFabLab) as a fablab unit dedicated to scientific research using digital fabrication with special focus on possible industrial applications. SciFabLab acts as a connecting hub between scientists, entrepreneurs and SMEs/large industry enabling efficient technology transfer i.e. putting scientific findings to practice by developing unique technology products. The article will present stages of SciFabLab establishment and conclusions about the ensuing stimulation of innovations and modernization of participating faculties, research centers and universities.[http://is.fink.rs/podaci/Milan_Matijevic/50/Stimulating%20innovations%20from%20university%20through%20the%20use%20of%20digital%20fabrication%20%E2%80%93%20case%20study.pdf

loop-mediated isothermal amplification (lamP) as a point-of-care SarS-covdetection method

Pravovremeno testiranje većeg broja ljudi na SARS-CoV-2 virus je povezano sa nižim mortalitetom od COVID-19 oboljenja. Međutim, većina zemalja nema mogućnosti za takvo masivno testiranje putem metode “PCR u realnom vremenu”, zbog visoke cene neophodne opreme i potrebe za stručnim osobljem. Zbog toga se razvijaju brze i ekonomičnije metode, koje se najčešće zasnivaju na izotermalnim metodama amplifikacije DNK. Za ove metode nije potreban ciklični termostat, zbog čega su primenljivije za terensku upotrebu. Fokus je na izotermalnoj amplifikaciji posredovanoj petljom (lamP), zbog njene visoke specifičnosti, mogućnosti za upotrebu neprečišćenih uzoraka i jednostavnosti merenja signala. Autori predstavljaju pregled najvažnijih radova o LAMP metodi za detekciju SARS-CoV-2 virusa objavljenih tokom 2020. godine, kao i opise nekoliko komercijalnih kompleta na bazi LAMP metode za COVID-19 testiranje.Massive testing for SARS-CoV-2 virus is related to lower mortality rates of COVID-19. Most countries face challenges to perform massive testing with the currently available methods (real-time PCR), due to expensive equipment and requirement of highly skilled personnel. To overcome these challenges, faster and cost-effective alternative detection methods are being developed, primarily based on isothermal methods of nucleic acid amplification (isoNAAs). PCR depends on precision instruments, high cleanliness of operating conditions and cannot be easily used on-site, while isoNAAs, which do not require thermal cyclers, are more applicable for point-of-care (PoC) use. Loop-mediated isothermal amplification (LAMP) is one of the isoNAA most in focus for COVID-19 tests due to its high specificity and possibility to use unpurified specimens in combination with simplified detection setup. The article gives a review of the most significant publications on use of LAMP for SARS-CoV-2 detection and of the several commercial LAMP-based COVID-19 testing kits.Ministarstva prosvete, nauke i tehnološkog razvoja Republike Srbije – broj ugovora 451-03-68/2020-14/ 200358 za autore M. Dj, T.K i Lj.J i broj ugovora 451-03-9/2021- 14/200125 za Ž.D.P. Autori zahvaljuju Jeleni Ognjenov na pomoći u pripremi ilustracija za rad

Point-of-Need DNA Testing for Detection of Foodborne Pathogenic Bacteria

Foodborne pathogenic bacteria present a crucial food safety issue. Conventional diagnostic methods are time-consuming and can be only performed on previously produced food. The advancing field of point-of-need diagnostic devices integrating molecular methods, biosensors, microfluidics, and nanomaterials offers new avenues for swift, low-cost detection of pathogens with high sensitivity and specificity. These analyses and screening of food items can be performed during all phases of production. This review presents major developments achieved in recent years in point-of-need diagnostics in land-based sector and sheds light on current challenges in achieving wider acceptance of portable devices in the food industry. Particular emphasis is placed on methods for testing nucleic acids, protocols for portable nucleic acid extraction and amplification, as well as on the means for low-cost detection and read-out signal amplification

Femtomolar detection of the heart failure biomarker NT-proBNP in artificial saliva using an immersible liquid-gated aptasensor with reduced graphene oxide

Measuring NT-proBNP biomarker is recommended for preliminary diagnostics of the heart failure. Recent studies suggest a possibility of early screening of biomarkers in saliva for non-invasive identification of cardiac diseases at the point-of-care. However, NT-proBNP concentrations in saliva can be thousand time lower than in blood plasma, going down to pg/mL level. To reach this level, we developed a label-free aptasensor based on a liquid-gated field effect transistor using a film of reduced graphene oxide monolayer (rGO-FET) with immobilized NT-proBNP specific aptamer. We found that, depending on ionic strength of tested solutions, there were different levels of correlation in responses of electrical parameters of the rGO-FET aptasensor, namely, the Dirac point shift and transconductance change. The correlation in response to NT-proBNP was high for 1.6 mM phosphate-buffered saline (PBS) and zero for 16 mM PBS in a wide range of analyte concentrations, varied from 1 fg/mL to 10 ng/mL. The effects of transconductance and Dirac point shift in PBS solutions of different concentrations are discussed. The biosensor exhibited a high sensitivity for both transconductance (2 uS/decade) and Dirac point shift (2.3 mV/decade) in diluted PBS with the linear range from 10 fg/mL to 1 pg/mL. The aptasensor performance has been also demonstrated in undiluted artificial saliva with the achieved limit of detection down to 41 fg/mL (~4.6 fM)

Recent advances on gradient hydrogels in biomimetic cartilage tissue engineering [version 1; referees: 2 approved]

Articular cartilage (AC) is a seemingly simple tissue that has only one type of constituting cell and no blood vessels and nerves. In the early days of tissue engineering, cartilage appeared to be an easy and promising target for reconstruction and this was especially motivating because of widespread AC pathologies such as osteoarthritis and frequent sports-induced injuries. However, AC has proven to be anything but simple. Recreating the varying properties of its zonal structure is a challenge that has not yet been fully answered. This caused the shift in tissue engineering strategies toward bioinspired or biomimetic approaches that attempt to mimic and simulate as much as possible the structure and function of the native tissues. Hydrogels, particularly gradient hydrogels, have shown great potential as components of the biomimetic engineering of the cartilaginous tissue

Uloga N-tipa voltazno zavisnih kalcijumskih kanala u degeneraciji aksona tokom eksperimentalnog autoimunog optickog neuritisa

Der entzündliche Befall des Sehnerven stellt eine der häufigsten klinischen Manifestationen der multiplen Sklerose (MS) dar, einer chronisch entzündlichen Erkrankung des zentralen Nervensystems (ZNS). In der Folge einer Optikusneuritis entwickeln etwa 30-50 % der Patienten eine bleibende Sehbehinderung verursacht durch eine Degeneration von Sehnerv-Axonen. In Vorarbeiten konnten wir zeigen, dass die experimentelle autoimmune Enzephalomyelitis (EAE), ausgelöst durch Immunisierung mit Myelin-Oligodendrozyten Glykoprotein (MOG), in braunen Norweger-(BN)-Ratten in 90 % der Fälle zu einer Optikusneuritis führt. Dieser Sehnervenbefall ist gekennzeichnet durch entzündliche Infiltration, Demyelinisierung und axonale Degeneration. Die genauen Mechanismen der entzündlich-bedingten axonalen Degeneration sind unklar. Es wird jedoch als wahrscheinlich angesehen, dass ein pathologischer Einstrom von Kalziumionen (Ca2+) in Axone eine wesentliche pathophysiologische Rolle spielt. Als mögliche Eintrittspforten für Ca2+ kommen dabei verschiedene Typen von spannungsabhängigen Kalziumkanälen (VDCC) in Betracht. Da Mangan (Mn2+) in Neurone ebenfalls über VDCC aufgenommen wird und eine Signalanhebung bei der Magnetresonanztomographie (MRT) bewirkt, haben wir die Technik des Mn2+-gestützten MRTs genutzt, um die Wirkung von Subtyp-spezifischen Kalziumkanalblockern zu untersuchen. Im Rahmen dieser Experimente führte die Applikation von ω Conotoxin GVIA, einem spezifischen Blocker von N-Typ Kalziumkanälen, zu einer signifikanten Hemmung der Mangan-Aufnahme in den entzündeten Sehnerven. Um parallel die Expression dieses Kanaltyps im Sehnerven zu untersuchen, haben wir immunhistochemische Färbungen für α1B durchgeführt. Dabei handelt es sich um die Untereinheit von N-Typ VDCC, die die Membranpore formt. Diese Färbungen ergaben signifikante Unterschiede in Lokalisation und Expression der Kanaluntereinheit in gesunden im Vergleich zu entzündlich befallenen Sehnerven. Während in gesunden, myelinisierten Sehnerven eine mäßige Expression von α1B nachzuweisen war, zeigte sich eine deutliche Hoch-Regulation in den Sehnerven der MOG-immunisierten Tiere. Ferner fand sich eine Korrelation zwischen der Expression von α1B und dem Ausmaß an Demyelinisierung, welches anhand von Myelin-spezifischen Färbungen quantifiziert wurde. Eine hoch-signifikante negative Korrelation ergab sich zwischen der Anzahl α1B-positiver Stellen im Sehnerven und der Anzahl überlebender Axone. In einem weiteren Schritt wurde dann die Wirksamkeit von ω Conotoxin GVIA mittels in vivo-Kalzium Imaging des Sehnerven überprüft. Dabei zeigte sich eine deutliche Reduktion des depolarisations-abhängigen Kalziumeinstroms in den entzündeten Sehnerven nach topischer Applikation des N-Typ Kalziumkanalblockers im Vergleich zu einer Kontrollgruppe Plazebo-behandelter Tiere. Die Applikation von -Conotoxin GVIA in gesunden Tieren hingegen führte lediglich zu einer leichten Verminderung des Kalziumeinstroms, der sich nicht signifikant von gesunden Kontrolltieren nach topischer Applikation von Kochsalzlösung unterschied. Diese Daten ergänzen die oben beschriebenen Expressionsanalysen des N-Typs spannungsabhängiger Kalziumkanäle und belegen ferner, dass die neu gebildeten Kanäle funktionsfähig sind. Zusammengefasst zeigen diese Ergebnisse, dass der pathologische Einstrom von Ca2+ im Rahmen der MOG-induzierten Optikusneuritis wesentlich über den N-Typ spannungsabhängiger Kalziumkanäle erfolgt. Die pathophysiologische Relevanz des gesteigerten Kalziumeinstroms über diesen Kanaltyp wurde ferner im Rahmen einer Therapiestudie mit ω-Conotoxin GVIA gezeigt. Dabei ergab die kontinuierliche zerebroventrikuläre Applikation des N-Typ-Kanalblockers ein signifikant vermindertes Maß an Demyelinisierung und axonalem Schaden. Unsere Daten einer ektopen Expression von N-Typ Kalziumkanälen im Sehnerven von BN- Ratten unter autoimmunen Bedingungen sowie die nachgewiesene Wirksamkeit von ω-Conotoxin GVIA lassen diese Substanz als mögliche neuroprotektive Therapie der autoimmunen Sehnerventzündung erscheinen

Microenvironment-Derived Stem Cell Plasticity

Plasticity is the hallmark of stem cells. At the same time, stem cells, like any other cell type, are influenced by their microenvironment and respond to it accordingly. A specific microenvironment is defined by a variety of factors, including biological and chemical factors, cell-cell interactions, but also metabolic and mechanical cues. Such dynamic and specialized microenvironment where the stem cells reside is considered a stem cell niche. Tissue injury as well as malignant tissue alterations lead to changes in the niche influencing the plasticity and biology of residing stem cells. Similarly, the niche changes upon tissue damage, which eventually induces differentiation of stem cells and ultimately regeneration of the tissue

Using Vertebrate Stem and Progenitor Cells for Cellular Agriculture, State-of-the-Art, Challenges, and Future Perspectives

Global food systems are under significant pressure to provide enough food, particularly protein-rich foods whose demand is on the rise in times of crisis and inflation, as presently existing due to post-COVID-19 pandemic effects and ongoing conflict in Ukraine and resulting in looming food insecurity, according to FAO. Cultivated meat (CM) and cultivated seafood (CS) are protein-rich alternatives for traditional meat and fish that are obtained via cellular agriculture (CA) i.e., tissue engineering for food applications. Stem and progenitor cells are the building blocks and starting point for any CA bioprocess. This review presents CA-relevant vertebrate cell types and procedures needed for their myogenic and adipogenic differentiation since muscle and fat tissue are the primary target tissues for CM/CS production. The review also describes existing challenges, such as a need for immortalized cell lines, or physical and biochemical parameters needed for enhanced meat/fat culture efficiency and ways to address them