Firdaningrum, Nimas Eka and Prof. Dr. dr. Kusworini Handono,, M. Kes., Sp.PK (K) and Dr. dr. Hani Susiati,, Sp.PK (K) (2023) Hubungan antara Penanda Immune Risk Profille Rasio Sel Limfosit T CD4/CD8 dengan Kadar sCD28 dan sCD86 pada Penuaan Sistem Imun. Magister thesis, Universitas Brawijaya.
Abstract
Aging (penuaan) merupakan suatu fenomena kompleks yang ditandai dengan perubahan serta penurunan progresif fungsi fisiologis dan homeostasis tubuh, yang menyebabkan meningkatnya kerentanan terhadap berbagai pajanan penyakit, proses degenerasi dan kematian. Proses penuaan ini dapat mempengaruhi berbagai sel, jaringan dan organ, termasuk sistem imun, disebut sebagai immune aging. Penuaan sistem imun merupakan proses alami pada individu tua, namun hal ini juga dapat terjadi pada pasien LES akibat immune exhaustion pada proses autoimun. Pada penuaan sistem imun mengakibatkan terjadinya disregulasi dan perubahan kompartemen sistem imun innate maupun adaptif. Saat ini IRP (immune risk profile), merupakan satu-satunya penanda prediktor penuaan sistem imun. Rasio sel limfosit T CD4/CD8 adalah IRP pertama sebelum ditemukannya penanda-penanda penuaan sistem imun lainnya, dan memiliki korelasi terbalik dengan risiko inflamasi dan/atau penyakit terkait usia. Namun IRP hanya dapat diamati dengan pemeriksaan flowsitometri yang cenderung tidak mudah, karena membutuhkan alat yang canggih dan keterampilan khusus. Oleh karena itu diperlukan studi lebih lanjut untuk meneliti penanda prediktif penuaan sistem imun dengan metode yang lebih efektif dan efisien. Molekul kostimulator CD28 dan CD86 merupakan pasangan reseptor-ligan yang berperan penting dalam regulasi sistem imun melalui sinyal kostimulator untuk aktivasi, proliferasi dan promosis kelangsungan hidup sel T. Kedua penanda ini merupakan salah satu penanda penting dalam penuaan sistem imun dan penyakit autoimun. Molekul kostimulator ini memiliki bentuk terlarut dalam serum (sCD28 dna sCD86) dan telah diteliti memiliki peran dalam berbagai penyakit autoimun, terbentuk akibat pelepasan dari sel membran atau produksi alternatif saat proses splicing mRNA. Penelitian ini bertujuan untuk mengetahui adanya hubungan antara penanda IRP rasio sel limfosit T CD4/CD8, baku emas deteksi immune aging dengan kadar sCD28 dan sCD86, sehingga diharapkan kedepannya kudua penanda terlarut ini dapat dikembangkan kit diagnostik immune aging. Penelitian ini merupakan studi kasus control, terdiri atas 87 responden penelitian, terbagi atas 40.2 % kelompok pasien LES, 36.8% lansia dan 23.0% individu muda. Rasio sel lifosit T CD4/CD8 didapatkan dari PBMC dan diukur dengan teknik flowcytometri. PBMC yang telah dicuci dengan PBS, disentrifugasi, kemudian pellet ditambahkan dengan cell staining buffer dan dilakukan staining PerCP anti human CD8 (Biolegend Katalog no. 344707) dan FITC anti human CD4 (Biolegend Katalog no. 300514). Pembacaan kuvet flowcytometri dilakukan dalam bentuk persentase sel, kemudian dihitung dengan BD Cell Quest software. Penanda sCD28 dan sCD86 pada serum diukur menggunakan metode ELISA. Kadar penanda kostimulator terlarut ini diukur dengan menggunakan Kit ELISA: Human Soluble CD28 (sCD28) ELISA Kit Mybiosource catalog number: MBS3801986, dan HSCD 86 (sCD86) ELISA Kit MBS catalog number: MBS702416. Pembacaan Optical Density pada 450 nm, dengan pembaca Microelisa Stripplate. Data penelitian ini dianalisis menggunakan software IBM SPSS versi 23, menggunakan uji komparasi Kruska Wallis untuk menentukan perbedaan rasio sel limfosit T CD4/CD8 serta kadar sCD28 dan sCD86 pada ketiga kelompok penelitian. Uji Korelasi Spearman dilakukan untuk mengetahui hubungan antara penanda IRP rasio sel limfosit T CD4/CD8 dengan kadar sCD28 dan sCD86.Pada penelitian ini didapatkan bahwa terdapat penurunan rasio sel sel limfosit T CD4/CD8 pada lansia dan individu muda, sedangkan terjadi peningkatan pada pasien LES. Kadar sCD28 dan sCD86 diamati mengalami peningkatan pada kedua kelompok penuaan imun, yaitu lansia dan pasien LES. Uji korelasi Spearman menunjukkan bahwa penanda IRP (immune risk profile) rasio sel limfosit T CD4/CD8 memiliki hubungan yang tidak signifikan dengan sCD28 (Kelompok individu ix muda, p value=0,995, r=-0,002; LES, p value=0,076, r=0,304; lansia, p value=0,101, r=-0,296) dan kadar sCD86 (Kelompok individu muda, p value=0,337, r=-0,227; LES, p value=0,080, r=-0,300; lansia, p value=0,485, r=-0,128) pada penuaan sistem imun. Hal ini menunjukkan bahwa semakin rendah rendah rasio sel limfosit T CD4/CD8 maka semakin meningkat pula kadar sCD28 dan sCD86, begitu pula sebaliknya. Hubungan antara IRP rasio sel limfosit T CD4/CD8 dengan kadar sCD28 dan sCD86 pada penuaan sistem imun dapat dijelaskan melalui keterkaitannya dengan peningkatan stress oksidatif dan proses inflamaging, pada saatnya akan mengakibatkan perubahan populasi dan diversitas sel limfosit T, berupa: penurunan rasio sel limfosit T CD4/CD8 dan peningkatan kadar penanda terlarut sCD28, serta peningkatan produksi sCD86 oleh APC. Sehingga dapat disimpulkan bahwa berdasarkan hasil penelitian ini penanda terlarut sCD28 dan sCD86 dapat dijadikan salah satu biomarker prediktif penuaan sistem imun di masa depan. Namun mekanisme dan perananan penanda terlarut sCD28 dan sCD86 terkait hubungannya dengan rasio sel limfosit T CD4/CD8 pada penuaan sistem imun masih perlu dikaji dan diteliti lebih lebih lanjut.
English Abstract
Aging known as a complex phenomenon, characterized by progressive changes and declines of physiological function and body homeostasis, lead to increased susceptibility of various disease, degenerative processes and death. Aging can affect various cells, tissues and organs, including the immune system, known as immune aging. Immune aging is a natural process in elderly, but it can also occur in same condition, such as SLE patients which undergo a premature immune aging due to immune exhaustion in autoimmune disease. Immune aging cause a dysregulation and changes in innate and adaptive immune systems compartments. Currently IRP (immune risk profile), is the only predictor marker of immune aging. The CD4/CD8 T lymphocyte ratio was the first IRP before other markers of immune aging were discovered. It has an inverse correlation with the risk of inflammation and/or age-related disease. However, IRP can only be observed by flow cytometry examination which tends to be difficult, because it requires sophisticated tools and special skills. Therefore, further studies are needed to examine predictive markers of immune system aging with more effective and efficient methods. Costimulator molecules CD28 and CD86 are receptor-ligand pairs that play an important role in immune system regulation through costimulator signals for activation, proliferation and promotion of T cell survival. These markers are important markers on detecting immune aging and autoimmune diseases. These costimulatory molecules have soluble forms in serum (sCD28 and sCD86) and have been investigated for their role in a variety of autoimmune diseases, produced either by membrane form shedding or alternative mRNA splicing. This study aims to determine the correlation between the IRP marker CD4/CD8 T lymphocyte ratio which is the gold standard for detecting the immune aging and the levels of sCD28 and sCD86 in immune aging, so that in the future these two soluble markers can be developed as the predictive biomarkers kit of the immune aging. It was an observational analysis study with case-control approach, 87 participants are divided into 40.2% SLE patient, 36.8% elderly and 23.0% health individuals. The CD4/CD8 T cell lymphocyte ratio was obtained from PBMC and detected using flowcytometry. PBMC that had been washed with PBS, centrifuged, then the pellet was added with cell staining buffer and stained with PerCP anti human CD8 (Biolegend Catalog no. 344707) and FITC anti human CD4 (Biolegend Catalog no. 300514). The flowcytometry cuvette was read in the form of cells percentage, calculated using the BD Cell Quest software. The sCD28 and sCD86 levels in serum were detected using ELISA Kit: Human Soluble CD28 (sCD28) ELISA Kit Mybiosource catalog number: MBS3801986, and HSCD 86 (sCD86) ELISA Kit MBS catalog number: MBS702416. The Optical Density was determined at 450 nm using a Microelisa Stripplate reader. IBM SPSS Statistics version 23 was used in this study analysis. The differences of CD4/CD8 T lymphocyte cells ratio and the levels of sCD28 and sCD86 in the three studies groups was conducted using ruska Wallis comparison test. The correlation between IRP markers CD4/CD8 T lymphocyte ratio and sCD28 and sCD86 levels was determined using Spearman Rank correlation test. This study shows that there was a decrease of CD4/CD8 T lymphocyte ratio in the elderly and young individuals, while there was increase in SLE patients. There were an increase levels of sCD28 and sCD86 observed in both immune aging groups, the elderly and SLE patients. Spearman correlation test showed there was an unsignificant and a weak negative correlation between IRP markers CD4/CD8 T lymphocyte ratio and the level of sCD28 (Young individuals, p value=0.995, r=-0.002; SLE, p value=0.076, r=0.304; elderly, p value=0.101, r=-0.296) and sCD86 levels (Young individuals, p value=0.337 , r=-0.227; SLE, p value=0.080, r=-0.300; elderly, p value=0.485, r=-0.128) in immune aging. This indicates that the lower the CD4/CD8 T lymphocyte ratio, xi the higher the sCD28 and sCD86 levels, and vice versa. The correlation between the ratio of CD4/CD8 T lymphocytes and levels of sCD28 and sCD86 in the aging immune system can be explained through its association with increase of oxidative stress and inflamaging processes in immune aging, which result in changes in the population and diversity of T lymphocyte cells; a decrease in CD4/CD8 T lymphocyte ratio and increase of the soluble marker sCD28 CD4/CD8 T lymphocyte ratio, as well as increase of sCD86 production by APC. Considering the results in this study, the soluble markers sCD28 and sCD86 can be used as immune aging predictive biomarkers in the future. However, the mechanism and role of the soluble markers sCD28 and sCD86 in correlation with CD4/CD8 T lymphocyte ratio in immune aging still need further study and research.
Other obstract
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| Item Type: | Thesis (Magister) |
|---|---|
| Identification Number: | 0422060171 |
| Subjects: | 600 Technology (Applied sciences) > 616 Diseases > 616.02 Special topics of disease > 616.025 Medical emergencies / Emergency medicine / Emergency nursing / Triage (Medicine) |
| Divisions: | S2/S3 > Magister Ilmu Biomedis, Fakultas Kedokteran |
| Depositing User: | Endang Susworini |
| Date Deposited: | 09 May 2023 01:13 |
| Last Modified: | 09 May 2023 01:13 |
| URI: | http://repository.ub.ac.id/id/eprint/198980 |
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