The text is a compendium of modern research techniques that enable the identification, separation and sequencing of proteins. They serve in the analysis of protein structure and also in the study of ...the relationship between protein structure and their biological function, including within metabolic processes. The richly illustrated publication is supplemented with additional materials.
For over 100 years the erythrocyte cell membrane attracted interest of transfusiologists mainly due to the antigens localized on their surface and associated risks of patient alloimmunisation and ...therefore the need of serological selection of donor's and recipient's blood. Presently it is known that RBC antigens and other membrane proteins play important transport and protective functions, and are involved in adhesion, maintenance of cell shape and in the process of aging and phagocytosis. Since the available results of retrospective clinical observations suggest an adverse effect of transfusion on selected groups of patients, it is important to undertake studies on the changes taking place within the cell membrane of erythrocytes stored in blood banks. Flow cytometric analysis of stored leucodepleted or non-leucodepleted erythrocyte concentrates, revealed significant changes in the level of expression of many RBC surface molecules: CD44, CD47, CD55, CD58, CD59, CD235a (GPA). In parallel, a significant development of proteomic analysis of stored RBCs is observed. Stored RBCs offer less variability of biological material, caused by drugs, illnesses, etc. when compared with clinical proteomics studies; however, the complexity of the methodology and the lack of uniform and comparable procedures may cause misinterpretation and even create artifacts. Still, modern research methods offer hope for the elaboration of aging biomarkers of stored RBCs as well as for raising the level of transfusion medicine quality.
Svrha: Željelo se pronaći optimalne uvjete i metode za identifikaciju maksimalnog broja proteina u uzorcima tkiva gingive pacijentice s agresivnim parodontitisom. Pritom smo se koristili tehnologijom ...Label-free kvantifikacije, tekućinske kromatografije i spektrometrije masa (LC –MS/MS), te usporedili proteom zdravih i bolesnih uzoraka tkiva gingive. Materijali i metode: Četiri uzorka gingivnog tkiva (dva zdrava i dva bolesna) uzeta su od pacijentice, inače nepušačice, koja boluje od teškog oblika generaliziranog agresivnog parodontitisa. Proteini tkivnog lizata razdvojeni su 1D gel-elektroforezom nakon koje je slijedila digestija u gelu te mjerenje proteina nanoljestvicom
HPLC-sistema preko nanoelektro spreja, ionizacijskog izvora i spektrometra mase. Dobiveni podaci obrađeni su u programu MaxQuant. Rezultati: Label-free kvantifikacija i LC-MS/MS analiza,
zajedno s pripremom uzoraka, 1D gel-elektroforezom i digestijom u gelu pokazali su se kao učinkovita metoda za analizu proteoma gingivnog tkiva. Ukupan broj identificiranih i kvantificiranih proteina iznosio je 2429. Proteini koji su pokazali pojačanu ekspresiju u tkivu zahvaćenom agresivnim parodontitisom bili su: imunoglobulinski gama-1 lanac regije C, imunoglobulinski kapa lanac regije C i imunoglobulinski gama-3 lanac regije C (Ig gamma-1 chain C region, Ig kappa chain C region i Ig gamma-3 chain C region), a desmoplakin, aneksini, proteini S100-A8/A9 i keratini pokazali su sniženu ekspresiju. Zaključak: Ovo eksperimentalno istraživanje omogućuje novi uvid u proteomski profil zdravog i bolesnog gingivnog tkiva, što bi moglo pridonijeti napretku u dijagnostici i prognozi te boljem razumijevanju patogeneze agresivnog parodontitisa. Tehnologija LCMS/ MS pokazala se učinkovitom kad je riječ o proteomskoj analizi gingivnog tkiva.
Proteomics was employed to identify the differentially expressed proteins between esophageal squamous cell carcinoma (ESCC) and adjacent normal esophageal tissues. ESCC tissues and adjacent normal ...tissues were obtained from 10 patients with ESCC and the proteins were extracted and subjected to two-dimensional gel electrophoresis (2-DE). The differentially expressed proteins were identified after image analysis, and matrix assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) was used to confirm these proteins. Immunohistochemistry was then performed to detect the expressions of HSP27 and ANX1 in ESCC tissues and adjacent normal tissues. A total of 6 differentially expressed proteins were identified by peptide mass fingerprinting, among which SCCA1, KRT4 and ANX1 were down-regulated and TIM1, MnSOD and HSP27 up-regulated in the ESCC. Immunohistochemistry showed HSP27 was highly expressed in the ESCC which, however, had a low expression of ANX1. These findings were consistent with those in proteomics. There were differentially expressed proteins between ESCC and adjacent normal tissues. The investigation of differentially expressed proteins between ESCC and normal esophageal tissue may provide evidence for the molecular pathogenesis of ESCC.
Uz pomoć proteomike, identifikovali smo proteine čija se ekspresija razlikuje u planocelularnom karcinomu jednjaka (PKJ) i susednim zdravim tkivima jednjaka. Uzorci tkiva PKJ i susednih normalnih tkiva uzeti su od 10 pacijenata sa PKJ a proteini su ekstrahovani i podvrgnuti dvodimenzionalnoj elektroforezi na gelu (2-DE). Proteini sa različitom ekspresijom su identifikovani posle analize slika, dok je za njihovo potvrđivanje upotrebljena tehnika MALDI-TOF-MS (matrix-assisted laser desorption ionisation time-of-flight mass spectrometry). Potom su obavljene imunohistohemijske analize kako bi se utvrdile ekspresije HSP27 i ANX1 u tkivima PKJ i susednim normalnim tkivima. Pomoću tehnike peptide mass fingerprintinga identifikovano je ukupno šest proteina sa različitim ekspresijama, od kojih je ekspresija SCCA1, KRT4 i ANX1 bila snižena, a TIM1, MnSOD i HSP21 povišena u PKJ. Imunohistohemijsko ispitivanje je pokazalo da je HSP27 veoma izražen u PKJ, u kom je, međutim, utvrđena niska ekspresija ANX1. Ovi rezultati se slažu sa onima dobijenim pomoću proteomike. Utvrđeno je da postoje proteini sa različitom ekspresijom u PKJ i susednim normalnim tkivima. Istraživanje različite ekspresije proteina u PKJ i normalnom tkivu jednjaka može doneti nove dokaze u vezi s molekulskom patogenezom PKJ.
Patogenezu složene i heterogene bolesti kao što je to rak dojke nije moguće u potpunosti
razjasniti klasičnim metodološkim pristupima koji su se koristi li posljednjih desetljeća
u molekularnoj ...medicini. Osobito je važno otkriti nove biljege koji bi pomogli u ranoj detekciji
bolesti te služili za razvoj ciljane terapije. Dosadašnji rezultati iz područja istraživanja raka
dojke u kombinaciji s globalnim metodama istraživanja statusa ekspresije gena i proteina
pokazali su kako je u patogenezu raka dojke uključen ljudski hormon rasta (hGH) te mehanizmi
autokrine regulacije preko proteina Pax-5. Također je identi fi ciran čitav niz novih potencijalnih
biomarkera kao što su to HOXA1, CHOP SH3GLB1, kazein kinaze, p53, aneksin XI, CDC25C, eIF-
4E i MAP kinaze 7, 14-3-3e, galekti n-1, aneksin-5, aneksin-1, LDH-B, GST-pi, akti n, vimenti n,
HSP70, CK18, moezin, SH3GLB1, SUB1, SND1 i TRIM28, osteoponti n i osteonekti n
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