Injectable Hydrogel Asam Hyaluronan (HA)-Polyethylene Glycol (PEG) Dengan Metode Crosslinking Enzimatik Untuk Terapi Penderita Degenerasi Diskus Intervertebralis

Chronic Low Back Pain (CLBP) atau nyeri punggung bawah merupakan salah satu masalah kesehatan yang sering dijumpai di masyarakat yang dapat mengenai siapa saja. Sekitar 60-80% dari seluruh penduduk dunia pernah mengalami nyeri pungung bawah, khususnya degenerasi diskus intervertebralis. Injectable hydrogel adalah cara terbaru untuk mengembalikan ketebalan diskus dan hidrasi yang ditimbulkan degenerasi diskus dengan minimal invasive surgery. Penelitian ini bertujuan mengetahui karakteristik injectable hydrogel berbasis Asam Hyaluronan (HA) dan Polyethylene Glycol (PEG), dengan crosslinker Enzim Horse Radish Peroxide Type I (HRP) dan mendapatkan komposisi enzim optimal pada injectable hydrogel sebagai terapi penderita degenerasi diskus intervertebralis. Karakterisasi dari penelitian ini yakni uji FTIR, uji swelling, uji degradasi, uji sitotoksisitas, dan uji in vitro injection model. Dari uji FTIR hidrogel menunjukkan adanya ikatan crosslinking pada sampel dengan penambahan enzim. Kemudian, hasil uji swelling menggunakan Phosphat Buffer Saline (PBS) yang mendekati nilai ideal untuk diskus intervertebralis adalah sampel dengan variasi konsentrasi enzim 0.25 μmol/menit/mL, yakni 33%. Uji degradasi membuktikan bahwa degradasi sampel akan semakin meningkat seiring dengan penurunan konsentrasi enzim HRP. Hasil dari uji sitotoksisitas melalui metode MTT Assay menunjukkan bahwa sampel menghasilkan prosentase sel hidup 90% dan bersifat tidak toksik. Pada uji in vitro injection model dibuktikan bahwa semakin tinggi konsentrasi enzim, keadaan gel ketika dilepas dari agarose akan semakin tidak rupture. HA-PEG-enzim HRP merupakan komposit polimer aman dan berpotensi diaplikasikan sebagai injectable hydrogel untuk degenerasi diskus intervertebralis berdasarkan gugus fungsi, derajat swelling, laju degradasi, sitotoksisitas, dan in vitro injection model. Kata Kunci : Injectable Hydrogel, HA, PEG, Enzim HR

Hyaluronic Acid (HA)-Polyethylene glycol (PEG) as injectable hydrogel for intervertebral disc degeneration patients therapy

Chronic Low Back Pain (CLBP) is one health problem that is often encountered in a community. Inject-able hydrogels are the newest way to restore the disc thickness and hydration caused by disc degeneration by means of minimally invasive surgery. Thus, polymers can be combined to improve the characteristic properties of inject-able hydrogels, leading to use of Hyaluronic Acid (a natural polymer) and Polyethylene glycol (PEG) with Horse Radish Peroxide (HRP) cross linker enzymes. The swelling test results, which approaches were the ideal disc values, were sampled with variation of enzyme concentrations of 0.25 µmol/min/mL. The enzyme concentrations were 33.95%. The degradation test proved that the sample degradation increased along with the decrease of the HRP enzyme concentration. The results of the cytotoxicity assay with MTT assay method showed that all samples resulted in the 90% of living cells are not toxic. In vitro injection, models demonstrated that higher concentration of the enzymes was less state of gel which would rupture when released from the agarose gel. The functional group characterization shows the cross linking bonding in sample with enzyme adding. The conclusion of this study is PEG-HAHRP enzyme are safe polymer composites which have a potential to be applied as an injectable hydrogel for intervertebral disc degeneration

Interleukin-13, Interleukin-10, Interferon-gamma and IDO Production in Response to Home Dust Mites in Allergic Asthma

BACKGROUND: Allergic asthma is a degenerative atopic disease caused by allergic or hypersensitivity type-1. More than 50% of people with allergic asthma are caused by the presence of house dust mites (HDMs) allergens.METHODS: The cellular immunity response was evaluated through a peripheral blood mononuclear cell (PBMC) culture isolated from blood, using the ficoll gradient technique. Subjects were atopic asthma groups and non-atopic asthma groups. PBMC from each subject cultured was stimulated with HDMs allergen, then incubated in a CO2 5% incubator, 37o C for 72 hours. With the multiplex assay method, interferon (IFN)-γ, interleukin (IL)-13 and IL-10 were measured, meanwhile indoleamine 2,3-dioxygenase level (IDO) was measured by the enzyme-linked immunosorbent assay (ELISA) sandwich methods.RESULTS: The IFN-γ production in the supernatant of PBMC cultures was stimulated by phytohemagglutinin (PHA), Roswell Park Memorial Institute (RPMI) medium and allergens. The IFN-γ production in allergen-stimulated supernatants showed higher level of IFN-γ in the nonatopic group (4,681,455±3,434,851) than atopic group (4,363,300±2,067,941) even though it was not statistically significant (p=0.078). There were no differences between the mean of IL-13 production in atopic asthma group and non-atopic group. The IL-10 production in allergenstimulated supernatants was shown to be higher in nonatopic group and were statistically significantly different (p=0.015). The IDO production in allergen-stimulated supernatants was shown to be higher in the non-atopic group (272,231±269,564) than in the actopic group (13,273±400), and it was significantly different (p=0.007).CONCLUSION: Cellular immune profile of subjects with allergic asthma to Dermatophagoides pterronyssinus (Der p) is characterized by a type-2 inflammatory response that is dominant compared to type-1 inflammation (higher IL-13 ratio compared to IFN-γ) and to the role of anti-inflammation (higher IL-13 ratio compared to IL-10). The decline in IDO production in allergic asthma subjects to Der p is thought to be related to the low cellular immune response in expressing IFN-γ compared to IL-13.KEYWORDS: interleukin-13, interleukin-10, IDO, PBMC, asthm