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Supplementary Material 1

Supplementary Material 2

Starostne spremembe imunskega sistema so vzrok za zmanjšano sposobnost obrambe proti okužbam, za pojav vnetnih bolezni in povečano incidenco raka. Obstaja več teorij, ki skušajo razložiti številne istočasne in prepletene procese staranja. Po teoriji staranja matičnih celic je za staranje tkiv in organov odgovoren upad sposobnosti matičnih celic, da bi uspešno obnavljale tkiva. Namen naše raziskave je bil na modelu laboratorijskih miši raziskati, ali lahko presaditev celic kostnega mozga (KM) mladih živali izboljša delovanje imunskega sistema starih živali ter s tem izboljša njihovo splošno zdravje in morda celo podaljša življenjsko dobo. Neobsevanim samicam BALB/c smo v 14., 16. in 18. (19.) mesecu starosti presadili celice KM mladih samcev. Vsem prejemnicam celic KM smo določili stopnjo himerizma v KM, v izoliranih populacijah limfocitov T, limfocitov B in nevtrofilcev ter v CFU kolonijah, pripravljenih iz KM prejemnic. Analizirali smo spremembe števila celic prirojenega in pridobljenega imunskega sistema ter funkcij imunskih celic po presaditvi (sposobnost endocitoze, citokinski odziv in test stimulacije proliferacije). Manjše število živali smo imunizirali s hemocianinom iz školjke (KLH) ter z analizo titra protiteles testirali njihov imunski odziv na antigen. Pri vseh miših smo redno ocenjevali 30 različnih parametrov ter določili indeks krhkosti. Spremljali smo njihovo življenjsko dobo ter ob smrti opravili raztelesbe ter beležili patoanatomske spremembe. Rezultati so pokazali, da so imele stare miši, ki so prejele celice mladega KM, izboljšane nekatere imunske parametre prirojenega in pridobljenega imunskega sistema v primerjavi s kontrolno skupino, vendar teh ni bilo dovolj, da bi lahko hipotezo raziskave potrdili. Prejemnice KM niso imele vidno boljšega splošnega zdravja, indeks krhkosti ni pokazal razlik med poskusno in kontrolno skupino in miši tudi niso živele dlje od kontrolne skupine miši. Age-related changes in immune functions are the cause of the increased vulnerability to infections, inflammatory diseases, and cancer incidence. There are many existing theories that try to explain the numerous, simultaneous, and interconnected aging processes. The stem cell theory of aging postulates that stem cells become inefficient at maintaining the original functions of the tissues. In our study, we used a mice model to observe whether non-conditioned bone marrow (BM) cell transplantation from young donors to old recipients would improve their immune system functions, improve their general health, and possibly even extend their lifespans. Female BALB/c mice received BM cells from young donors at 14, 16 and 18(19) months of age. We determined chimerism levels in the BM, isolated T cell, B cell and neutrophil populations, and in colony-forming units (CFU) in BM. We analysed the changes of relative cell number of the innate and adaptive immune systems, and conducted functional assays (endocytic capability, cytokine response, and cell proliferation). We immunized a small group of mice with keyhole limpet haemocyanin (KLH) antigen and tested their immune response to antigen. All mice were regularly assessed using 30 parameters to calculate a frailty index (FI). We noted their lifespans, conducted post-mortem examinations, and recorded any pathoanatomical changes. Our results showed that old mice, transplanted with the BM cells of young donors, had some improvement in the immune parameters of their innate and adaptive systems, however, the evidence was insufficient for us to confirm our hypothesis. Recipient mice did not experience a noticeable improvement in their general health, and the FI showed no significant differences between the transplanted and control mice. In addition, the transplanted group did not live longer than the control group.

Staranje je zapleten, več faktorski proces, ki prizadane celice, tkiva in organe. S staranjem organizma se stara tudi imunski sistem, ki je sicer ključen za obrambo organizma pred škodljivim okoljem. S starostjo povezane okvare imunskega sistema vodijo v nastanek večine s starostjo povezanih bolezni kot so rak, avtoimunske bolezni in srčno-žilne bolezni ter povročijo povečano dovzetnost posameznika za okužbe. Ena izmed možnih terapij, ki bi lahko izboljšala delovanje imunskega sistema v starosti, je presaditev mladih krvotvornih matičnih in progenitorskih celic (KMPC), ki bi jih posamezniku odvzeli in shranili v njegovi mladosti ter ponovno presadili v njegovi starosti. Sklepamo, da bi na ta način mlade in zdrave KMPC po presaditvi »pomladile« stari imunski sistem ter izboljšale njegovo delovanje, ob enem pa bi posamezniku podaljšali njegovo življensko dobo. Omenjeni princip se imunuje heterohrona avtologna presaditev mladih krvotvornih matičnih celic. V raziskavi smo v mišjem modelu uporabili nov način presaditve KMPC s predhodno mobilizacijo endogenih KMC v periferno kri. V ta namen smo uporabili molekulo CASIN, ki začasno izprazni nišo kostnega mozga prejemnika, na ta način pa omogoči večjemu številu presajenih celic pritrjanje vanjo. Z uporabo molekule CASIN smo po presaditvi mladih KMPC starim prejemnicam dosegli visok odstotek donorskega himerizma, dokazali nekatere izboljšave v imunskem sistemu in rahlo povišali mediano preživetja. Kljub temu pa bi bilo za prenos v humano medicino potrebno izvesti dodatne raziskave, ki bi potrdili ugodne učinke tovrstne terapije. Aging is a complex multi-factorial process that affects cells, tissues and organs. It also affects the immune system, which is crucial for the body's defense against the harmful environment. Age-related defects of the immune system lead to the development of most age-related diseases, such as cancer, autoimmune and cardiovascular diseases and cause an increased susceptibility to infections. One of the strategies that could improve the function of the immune system of an old individual is the transplantation of young hematopoietic stem and progenitor cells (HSCs) which would be collected from an individual in his/her youth and cryopreserved for a long time. After re-infusion later in life of the same individual, young and healthy HSPCs could »rejuvenate« the old immune system and improve its functioning. Furthermore, individuals lifespan could be prolonged. The process is called heterochronous autologous hematopoietic stem cell transplantation. In this study we used a new method of HSC transplantation with prior mobilization of endogenous HSCs into peripheral blood. For this purpose, we used the molecule CASIN, which temporarily empties the recipient's niche, thus allowing engraftment of a larger number of transplanted cells. With the use of the CASIN molecule, we achieved a high percentage of donor chimerism after transplantation of young HSPCs into old recipients. In the recipients of young HSPCs, we detected some improvements in the function of the immune system and a slighty higher median lifespan. Nevertheless, for translation into human medicine, additional research would be needed to confirm the beneficial effects of this type of therapy.