Ny mærkningstilgang gør det muligt at undersøge pakker, celler udsendes for at få indsigt i sundhed

Vores celler kommunikerer konstant, og forskerne har udviklet en effektiv måde at finde ud af, hvilke beskeder de sender i proteinfyldte biologiske kufferter kaldet exosomer.

Disse sfæriske exosomer, som ligger i den indre membran af en celle, men til sidst vil gå ud for at komme ind i en anden celle, transporterer store molekyler som proteiner, en grundlæggende byggesten i kroppen og drivere til biologisk aktivitet, og RNA, som producerer protein.

"Dette er en løbende proces," siger Dr. Sang-Ho Kwon, cellebiolog i afdelingen for cellulær biologi og anatomi ved Medical College of Georgia på Augusta University, og der er stigende beviser for, at det forekommer både i sundhedstilstande og sygdom.

"Vi forsøger at finde ud af dette puslespil om, hvad exosomer gør i forskellige scenarier," siger Kwon. Han er tilsvarende forfatter til en undersøgelse i Journal of Extracellular Vesicles beskriver en mærkningsteknik, som han og hans forskerhold har udviklet til at analysere indholdet af exosomer fra enhver specifik celletype for bedre at forstå deres rolle i velvære og sygdom.

"Deres indhold kan hjælpe med at fortælle os, hvad vores celler fortæller hinanden," siger Kwon og giver sandsynligvis tidlige spor om, at vi bliver syge og hjælper os med bedre at forstå, hvordan vi bliver syge.

Det menes, at gods bliver læsset tidligt i dannelsen af ​​exosomer af deres forløber-endosomer, nær cellemembranen, som fungerer meget som at fylde postbilen på postkontoret, før den begiver sig ud på sin rute. Exosomer vil blive der, indtil de frigives af cellen for at rejse til andre celler.

Kwon og hans team ønskede at fange lasten tidligt i processen.

Lige nu er den vigtigste måde at studere exosomindhold på først at tage exosomer ud af kontekst, for at isolere dem, en ret besværlig proces, der kan give inkonsistente resultater. Faktisk kan det isolere en anden type vesikler, dybest set biologiske rum i vores krop, hvoraf exosomer kun er én type.

MCG-teamet har udviklet en mere effektiv metode, der gør det muligt kun at studere indholdet af exosomer og studere, hvor de er.

Deres mærkningssystem inkluderer en variant af APEX eller ascorbatperoxidase, som er fusioneret til et andet protein, der vides at opsøge exosomer. "APEX er en slags missil, der får mig indenfor," siger Kwon. APEX har en høj affinitet for biotin, et B-vitamin, som binder sig til nærliggende proteiner, som dem, det udviklende exosom bærer, mærker dem og hjælper med at identificere dem. Biotin kan også passere gennem cellemembranen, som exosomer er bagved. Endnu et protein, streptavidin, som binder naturligt til biotin, sætter dem i stand til at oprense og klart identificere proteinlasten såvel som det RNA, der vil producere fremtidige proteiner, ved hjælp af analyse leveret af massespektrometri.

Kwons fokus er nyreskade, og de har brugt deres system til at vise, at oxidativt stress, et biprodukt af brugen af ​​ilt, som er overdreven og ødelæggende i sygdomstilstande, ændrer lastindholdet i exosomer, der dannes af nyreceller og findes i urinen. . For eksempel ændrede ekspressionsniveauer af nogle proteiner sig, og nogle proteiner forsvandt endda.

Their technique should ease development of databases of the usual content of a variety of different cell types that will enable comparative studies of what happens to their content in different disease states like the kidney injuries Kwon studies, or cancer.

"It turns out that by looking at the exosomes in the urine or blood, and by looking at what is inside, we can tell whether the cell is injured or a healthy cell," he says.

Their first use of the labeling system was in live kidney cells in culture. They now want to use it in an animal model of kidney disease.

The scientific team says the labeling system additionally can help trace how exosome content changes over time and potentially how cells are responding to treatment in the case of disease.

Exosomes are known to play a key role in cell communication, both between cells of the same type and with other types. Again, there is increasing evidence of the role exosomes play in disease, including sharing with other cells the news that they are sick and potentially even helping spread disease. "It's not just passing good news. it also passes bad news," Kwon says.

He notes their cargo no doubt varies in those diverse scenarios, an important reason to be able to detect what exosomes are carrying. Changes may ultimately serve as good way to monitor response to treatment, another aspect of exosome research that is "exploding," Kwon says. Scientists also are exploring the potential of using exosomes to actually deliver treatment, by filling these biological packages with medication that can be delivered directly to the desired location.

In fact, immune cells, which are pivotal in health and disease, also are releasing exosomes. These biological compartments also appear to play an important role in taking cellular debris and other trash out of the cells.

"It's an emerging field, right now," says Kwon. Proteins are the primary occupant because they can send signals, but they can also bind to other proteins and change their function, he says. RNA can do the same, and tiny microRNA can alter gene expression and consequently cell function.

Kwon's interest in exosomes was sealed when, as a postdoc at the University of California San Francisco, he grew kidney tubules, which return vital nutrients to the blood and eliminate undesirables in the urine, in a dish and found evidence that exosomes were playing a key role in the changing gene dynamics there.

He calls the focus on exosomes "reverse science," with most people looking at how the cell changes while he and a growing number of colleagues are looking at the packages the cell is sending out to understand what the cell is up to. While it may not seem like it to most people, he says it's actually a less complex way to approach cell activity because you are looking at a smaller package with far fewer proteins. + Udforsk yderligere

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