Hasil Pencarian  ::  Simpan CSV :: Kembali

"Latar belakang: Terapi sel punca dikembangkan sebagai alternatif terapi gagal jantung akibat infark miokardium. Bermacam tipe sel dengan berbagai metode implantasi telah banyak dikembangkan tetapi belum mendapatkan hasil optimal. Sel h-AECs (Human Amnion Epithelial Stem Cells) memiliki sifat yang sangat mendukung sebagai sumber sel bagi terapi sel punca pada jantung. Teknologi rekayasa jaringan dengan melakukan ko-kultur kardiomiosit dan h-AECs pada biomaterial scaffold diyakini dapat menjawab permasalahan pada pengembangan terapi sel punca pada gagal jantung.Metode: Penelitian ini adalah studi eksperimental in-vitro dengan penyemaian ko-kultur sel kardiomiosit dan h-AECs ke dalam scaffold patch. Kardiomiosit berasal dari otot ventrikel kanan pasien penderita penyakit tetralogy of Fallots yang dilakukan operasi koreksi TOF. Sedangkan sel h-AECs didapat dari epitel amnion yang merupakan limbah operasi seksio sesarea. Setelah dilakukan karakterisasi pada kardiomiosit dan h-AECs, dilakukan ko-kultur pada scaffold amnion dengan perbandingan densitas penyemaian 1:5 dan 1:6. Evaluasi hasil ko-kultur dilakukan dengan penilaian viabilitas sel, ekspresi gen spesifik kardiomiosit dan uji toksisitas patch.Hasil: Hasil ko-kultur kardiomiosit dan h-AECs tidak terdapat perbedaan bermakna pada rerata jumlah sel viabel pada hari kedua dan kelima (p>0,05). Sedangkan pada hari kedelapan terdapat perbedaan bermakna pada jumlah sel viabel, rasio 1:5 menghasilkan jumlah sel viabel lebih baik dibanding rasio 1:6 (p=0,011). Ekspresi gen spesifik kardiomiosit konsisten tampak pada kelompok rasio 1:6 dan mulai menunjukkan signifikan pada hari kedelapan, terdapat perbedaan bermakna pada ekspresi gen di hari kedelapan, kelompok rasio 1:6 mengekspresikan gen cTnT dan ACTN2 lebih baik dibanding kelompok 1:5 (p=0,000 dan p=0,001). Pada uji toksisitas, tidak terdapat perbedaan bermakna pada jumlah ATP dan kadar TNFα antara kelompok 1:5 dan 1:6.Simpulan: Teknik ko-kultur yang dikembangkan dapat menghasilkan sel kardiomiosit baru. Kelompok rasio 1:6 menghasilkan sel yang memiliki sifat spesifik kardiomiosit lebih baik dibanding kelompok rasio 1:5 tetapi menghasilkan jumlah sel viabel lebih sedikit. Patch hasil ko-kultur tidak bersifat toksik.

---

Background: Stem cell therapy was developed as an alternative therapy for heart failure due to myocardial infarction. Various types of cells with various methods of implantation have been developed but have not yet obtained optimal results. h-AECs (Human Amnion Epithelial Stem Cells) have very supportive properties as a source of cells for stem cell therapy in the heart. Tissue engineering technology by co-culturing cardiomyocytes and h-AECs on scaffold biomaterials is believed to be able to answer problems in the development of stem cell therapy in heart failure.

Method: This study is an in-vitro experimental study by seeding co-cultures of cardiomyocytes and h-AECs into the scaffold patch. Cardiomyocytes were derived from the right ventricular muscle of patients with tetralogy of Fallot disease who underwent TOF correction surgery. Meanwhile, the h-AECs cells were obtained from the amniotic epithelium which is the waste from cesarean section. After characterization of cardiomyocytes and h-AECs, co-culture was performed on amnion scaffold with seeding density ratio 1:5 and 1:6. Evaluation of co-culture results was carried out by assessing cells viability, expression of specific cardiomyocytes gen and patch toxicity tests.

Result: The results of co-culture of cardiomyocytes and h-AECs showed no significant difference in the mean number of viable cells on the second and fifth days (p>0.05). While on the eighth day there was a significant difference in the number of viable cells, a ratio of 1:5 resulted in a better number of viable cells than a ratio of 1:6 (p=0.011). Cardiomyocyte-specific gene expression was consistently seen in the 1:6 ratio group and began to show significantly on the eighth day, there was a significant difference in gene expression on the eighth day, the 1:6 ratio group expressed cTnT and ACTN2 genes better than the 1:5 group (p= 0.000 and p=0.001). In the toxicity test, there was no significant difference in the amount of ATP and TNFα levels between the 1:5 and 1:6 groups.

Conclusion: The developed co-culture technique can generate new cardiomyocytes. The 1:6 ratio group produced cells that had better cardiomyocyte-specific properties than the 1:5 ratio group but produced fewer cells. Co-culture of h-AECs and cardiomyocytes on patch was not toxic."

"Infark miokard menyebabkan kematian kardiomiosit dan remodeling jantung pada situasi patologis. Pascainfark jantung tidak mampu mengatasi kehilangan kardiomiosit meskipun telah dilakukan rekanalisasi atau revaskularisasi. Oleh karena itu, diperlukan metode untuk mengembalikan fungsi jantung. Sel punca dapat memperbaharui diri dan berdiferensiasi menjadi berbagai tipe sel namun kesintasannya pada pasien masih rendah. Untuk meningkatkan retensi dan regenerasi sel punca di miokardium dapat digunakan perancah/scaffold dan sistem ko-kultur, namun belum ada penelitian tentang hal tersebut. Penelitian ini bertujuan mengembangkan terapi infark menggunakan injeksi hidrogel transepikardial dan implantasi di epikardial perancah patch membran amnion yang dideselularisasi menggunakan amniotic epithelial cells (AEC) dengan ko-kultur kardiomiosit. Penelitian ini menggunakan post-test only control group design yang dilakukan di Institut Pertanian Bogor dan Fakultas Kedokteran Universitas Indonesia dari Juli 2021–Oktober 2022. Subjek penelitian adalah 15 babi Sus scrofa domesticus usia 2-3 bulan dibagi tiga kelompok: pAEC, pAEC + kardiomiosit, kontrol positif, dan 1 babi sebagai kontrol negatif. Torakotomi dilakukan untuk membuat model infark dengan ligasi arteri proximal branch to left ventricle (PLV) dilanjutkan implantasi pAEC dengan atau tanpa ko-kultur kardiomiosit pada kelompok terapi, kemudian diobservasi selama 6–8 minggu. Luas infark diukur dengan late gadolinium enhancement MRI; remodeling ventrikel kiri dengan ekokardiografi untuk menilai kontraktilitas, fibrosis dengan IHK, kardiomiogenesis dan regulasi apoptosis dengan RT-PCR, angiogenesis dinilai dengan IHK, dan fraksi ejeksi dinilai dengan ekokardiografi. Luas infark menurun pada kedua kelompok terapi (2,5 [2,00–3,00]% dan 3,60 ± 1,34% vs 9,50 ± 1,91%). Pewarnaan HE dan Masson trichrome menunjukkan berkurangnya proses fibrosis pada kedua kelompok, dikonfirmasi dengan hiperekspresi kolagen1 yang padat dan kaku pada kontrol positif dibandingkan kedua kelompok terapi yang memiliki ekspresi kolagen3 lebih dominan. Ekspresi α-smooth muscle actin pada kedua kelompok tampak tersebar menunjukkan penurunan fibrosis dan kontrol positif menunjukkan peningkatan fibrosis. Peningkatan kardiomiogenesis pada kedua kelompok dikonfirmasi dengan peningkatan ekspresi gen cardiac troponin T, gen myosin heavy chain, gen Nkx.2.5, gen c-Kit, dan penanda otot fungsional α-actinin. Penurunan apoptosis dikonfirmasi dengan penurunan ekspresi gen modulator apoptosis p21 dan ekspresi gen p53 yang berarti diferensiasi sel punca tidak bersifat tumorigenik. Regulasi apoptosis melalui ekspresi kaspase-9 tidak berbeda bermakna. Peningkatan angiogenesis dikonfirmasi dengan peningkatan ekspresi von Willebrand Factor dan ekspresi α-smooth muscle actin yang tersebar. Ekokardiografi menunjukkan perbaikan regional wall motion abnormality lebih banyak pada kelompok terapi daripada kontrol positif dan fraksi ejeksi tidak berbeda bermakna antar kelompok. Disimpulkan kombinasi injeksi hidrogel transepikardial dan implantasi di epikardial perancah patch membran amnion yang dideselularisasi dengan ko-kultur AEC dan kardiomiosit dapat mengurangi luas infark dan remodelling ventrikel kiri, serta meningkatkan angiogenesis pada babi model infark.

---

Myocardial infarction induces cardiomyocyte death and remodelling a pathological condition. The post-infarct heart is unable to deal with cardiomyocyte loss despite recanalization or revascularization. Therefore, a procedure is required to restore cardiac function. Stem cells can self-renew and specialize into multiple cell types however the survival of stem cells in patients is still poor. To promote the retention and regeneration of stem cells in the myocardium, scaffolds and co-culture systems may be applicated, although there are no study findings on this issue. This study aimed to develop myocardial infarction therapy using transepicardial hydrogel injection and epicardial decellularized amniotic membrane scaffold patch implantation using amniotic epithelial cells (AEC) with cardiomyocyte co-culture. This study used a post-test-only control group design performed at the IPB University and the Faculty of Medicine, Universitas Indonesia, from July 2021 to October 2022. The study subjects were 15 Sus scrofa domesticus pigs aged 2-3 months placed into three groups: pAEC, pAEC + cardiomyocytes, positive control, and 1 pig as a negative control. Thoracotomy was conducted to create an infarct model with the proximal branch to left ventricle (PLV) artery occlusion followed by pAEC implantation with or without cardiomyocyte co-culture in the therapy group, then evaluated for 6–8 weeks. Infarct size was determined by late gadolinium enhancement MRI, left ventricular remodeling by echocardiography to evaluate contractility, fibrosis by IHC, cardiomyogenesis and regulation of apoptosis by RT-PCR, angiogenesis was assessed by IHC, and ejection fraction by echocardiography. Infarct size reduced in both therapy groups (2,5 [2,00–3,00]% and 3,60 ± 1,34% vs 9,50 ± 1,91%). HE and Masson trichrome staining demonstrated decreased fibrosis in both groups, confirmed by hyperexpression of dense and stiff collagen 1 in the positive control compared to the two therapy groups with more dominant collagen 3 expressions. The α-smooth muscle actin expression in both groups seemed to be scattered suggesting reduced fibrosis while the positive control showed increased fibrosis. Increased cardiomyogenesis in both groups was confirmed by increased expression of the cardiac troponin T gene, the myosin heavy chain gene, the Nkx.2.5 gene, the c-Kit gene, and the functional muscle marker α-actinin. The reduction in apoptosis has been confirmed by lower expression of the p21 apoptosis modulator gene and p53 gene expression, which suggests that stem cell differentiation is not tumorigenic. The control of apoptosis by caspase-9 expression was not significantly different. Increased angiogenesis was verified by increased von Willebrand Factor expression and scattered expression of α-smooth muscle actin. Echocardiography showed greater improvement in regional wall motion abnormalities in the therapy groups than in the positive control, and the ejection fraction was not significantly different between groups. It was concluded that the combination of transepicardial hydrogel injection and epicardial decellularized amniotic membrane scaffold patch implantation using AEC with cardiomyocyte co-culture could reduce infarct size and left ventricular remodeling, as well as increase angiogenesis in infarct model pigs."

"Isolasi dan ekspansi sel punca kanker payudara secara in vitro tanpa menghilangkan sifat pluripotensi sel sangat diperlukan untuk mempelajari karakteristik sel punca kanker payudara. Penelitian ini bertujuan untuk mencari suatu kondisi kultur terbaik dengan sistem ko-kultur untuk mempertahankan sifat pluripotensi. Sel tersebut diko-kultur dengan feeder layer yang terbentuk dari Mouse Embryonic Fibroblast (MEF). Pengukuran ekspresi penanda permukaan CD44+/CD24-menggunakan metode spektrofluorometri, sedangkan pengukuran ekspresi gen SOX2 dilakukan secara kuantitatif dengan metode real time polymerase chain reaction. Pada pengukuran ekspresi gen SOX2 dilakukan normalisasi menggunakan house keeping gene PUM1 sebagai kontrol dalam. Hasil penelitian menunjukkan bahwa ekspresi CD44+/CD24-pada sel punca kanker payudara yang diko-kultur dengan MEFs menggunakan media CM dan DMEM lebih tinggi dibandingkan dengan yang tidak diko-kultur dengan MEF menggunakan CM dan DMEM. Ekspresi gen SOX2 meningkat pada sel yang diko-kultur dengan MEF di CM dan sel yang diko-kultur dengan MEF dalam DMEM yaitu berturut-turut 83,28 dan 48,33 kali dari kontrol masing masing. Hasil penelitian menunjukan bahwa kondisi kultur terbaik untuk mempertahankan pluripotensi sel punca kanker payudara adalah ko-kultur dengan MEF menggunakan CM sebagai media.

---

In objective to understand the role and importance of breast cancer stem cells and the molecular and cellular events that occur during cancer development, it is essential to isolate and propagate breast cancer stem cells in long-term in vitro culture without loosing their pluripotency. This study aimed to find the best culture condition with co-cultured system to maintain the breast cancer stem cells pluripotency. The cell cultured on top of a feeder layer formed by mouse embryonic fibroblast (MEF). We observed the expression of breast stem cell markers CD44+/CD24-using spectrofluorometry and analyzed the expression of gene SOX2 using quantitative real-time polymerase chain reaction (qRT-PCR).

Fluorescence spectroscopy results showed that CD44+/CD24-expression of cancer stem cells co-cultured in CM and DMEM high glucose with MEF is higher than one cultured in CM or in DMEM high glucose without MEF only. After being normalized by using house keeping gene, PUM1, SOX2 gene expression levels in cells cultured in CM and DMEM with MEF i.e 83,28 and 48,33 times higher, respectively, compared to their control. Result showed that the best condition to maintain pluripotency of breast cancer stem cells was to co-culture the cells with MEF using CM as medium."

"Sel punca mesenkim (MSC) dan sel punca pluripoten terinduksi (iPSC) telah dilaporkan mampu berdiferensiasi menjadi hepatosit secara in vitro dengan berbagai tingkat maturasi hepatosit. Sebuah metode sederhana untuk proses deselulerisasi perancah hati telah dikembangkan oleh Fakultas Kedokteran Universitas Indonesia. Penelitian ini bertujuan untuk mengevaluasi diferensiasi hepatosit dari iPSC dibandingkan dengan MSC dalam perancah hati yang dideselularisasi. Langkah pada penelitian ini adalah mengkultur iPSC dan MSC, mendeselularisasi hati kelinci, menyemai kultur sel ke dalam perancah, dan mendiferensiasikan menjadi hepatosit selama 21 hari dengan protokol Blackford yang dimodifikasi. Pemeriksaan dilakukan dengan pewarnaan Haematoxylin Eosin (HE), Masson Trichrome (MT), imunohistokimia (IHK) albumin dan cytochrome 3A4 (CYP3A4). Ekspresi gen albumin, cytochrome P450 (CYP450), dan cytokeratin-19 (CK-19) dianalisis menggunakan qRT-PCR. Pemeriksaan scanning electron microscope (SEM) dan immunofluorescence (IF) marker hepatocyte nuclear factor 4 alpha (HNF4-α) dan CCAAT/enhancer-binding protein alpha (CEBPA) dilakukan. Diferensiasi hepatosit dari iPSC dalam perancah hati yang dideselulerisasi dibandingkan dengan diferensiasi hepatosit dari MSC dalam perancah hati yang dideselulerisasi menunjukkan pembentukan sel tunggal dan kapasitas adhesi pada perancah yang lebih sedikit, dan penurunan tren ekspresi albumin dan CYP450 yang lebih rendah. Jumlah penyemaian sel awal yang lebih rendah menyebabkan hanya beberapa iPSC menempel pada bagian-bagian tertentu dari perancah hati yang dideselularisasi. Injeksi jarum suntik manual untuk reselulerisasi yang tidak merata menciptakan pola pembentukan sel tunggal oleh hepatosit dari diferensiasi iPSC di perancah hati yang dideselulerisasi. Hepatosit dari diferensiasi MSC memiliki kapasitas adhesi lebih tinggi ke perancah hati yang dideselulerisasi yang mengarah pada peningkatan tren ekspresi albumin dan CYP450. Penurunan ekspresi gen CK-19 lebih banyak terjadi pada diferensiasi hepatosit dari iPSC. Hasil tersebut dikonfirmasi oleh adanya sinyal positif protein HNF4-α dan CEBPA dengan pemeriksaan IF yang menunjukkan hepatosit yang dewasa. Kesimpulan dari penelitian ini adalah diferensiasi hepatosit dari iPSC pada perancah hati yang dideselularisasi lebih dewasa dengan adhesi sel-matriks ekstraseluler lebih rendah, distribusi sel spasial saling berjauhan, dan ekspresi albumin dan CYP450 lebih rendah dibandingkan dengan diferensiasi hepatosit dari MSC pada perancah hati yang dideselularisasi.

---

Mesenchymal stem cells (MSC) and induced pluripotent stem cells (iPSC) have been reported able to differentiate to hepatocyte in vitro with varying degree of hepatocyte maturation. A simple method to decellularized liver scaffold has been established by Faculty of medicine Universitas Indonesia.

This study aims to evaluate hepatocyte differentiation from iPSCs compared to MSCs in decellularized liver scaffold. iPSCs and MSCs were cultured, rabbit liver were decellularized, cell cultures were seeded into the scaffold, and differentiated into hepatocytes for 21 days with modified Blackford protocol. Haematoxylin-Eosin (HE), Masson Trichrome (MT), immunohistochemistry (IHC) albumin and CYP3A4 was performed. Expression of albumin, cytochrome P450 (CYP450) and cytokeratin-19 (CK-19) genes were analyzed using qRT-PCR. Scanning electron microscope (SEM) and immunofluorescence (IF) examination of hepatocyte nuclear factor 4 alpha (HNF4-α) and CCAAT/enhancer-binding protein alpha (CEBPA) marker was performed.

Hepatocyte differentiated iPSCs compared with hepatocyte differentiated MSCs in decellularized liver scaffold single–cell–formation and lower adhesion capacity in scaffold, and decrease trends of albumin and CYP450 expression. Lower initial seeding cell number causes only a few iPSCs to attach to certain parts of decellularized liver scaffold. Manual syringe injection for recellularization abruptly and unevenly create pattern of single–cell–formation by hepatocyte differentiated iPSCs in the decellularized liver scaffold. Hepatocyte differentiated MSCs have higher adhesion capacity to decellularized liver scaffold that lead to increase trends of albumin and CYP450 expression. CK-19 expression gene diminished more prominent in hepatocyte differentiated iPSCs.

These results were confirmed by the presence of HNF4-α and CEBPA positive signal protein with IF examination, showing mature hepatocyte.The conclusion of this study is hepatocyte differentiated iPSCs in decellularized liver scaffold differentiation is more mature with lower cell-extracelullar matrix adhesion, spatial cell distribution far from each other, and lower albumin and CYP450 expression than hepatocyte differentiated MSCs in decellularized liver scaffold."

"Latar belakang: Penyakit jantung koroner merupakan salah satu penyebab utama kematian di negara berkembang, termasuk Indonesia. Penyakit ini merupakan salah satu penyebab gagal jantung. Terapi yang selama ini dilakukan belum sepenuhnya mampu memperbaiki kerusakan otot jantung yang telah terjadi. Terapi sel punca baik injeksi maupun patch juga masih belum memperlihatkan hasil yang memuaskan. Penggunaan patch jantung dihadapkan pada masalah seperti rendahnya viabilitas sel yang dihasilkan. Hipotermia pada suhu 4°C yang digunakan untuk isolasi kardiomiosit selama ini dikaitkan dengan gangguan aktivitas sel yang menyebabkan penurunan jumlah sel viabel yang dihasilkan. Suhu 37°C yang merupakan suhu fisiologis tubuh dinilai mampu menghasilkan viabilitas sel lebih baik. Metode: Studi ekperimental in vitro dilakukan di Pelayanan Jantung Terpadu Rumah Sakit Cipto Mangunkusumo (PJT-RSCM) dan Indonesian Medical Education and Research Institute (IMERI) Fakultas Kedokteran Universitas Indonesia dalam periode Desember 2019 hingga November 2020 dengan mengikutsertakan subjek pasien yang menjalani operasi total koreksi tetralogy of Fallot. Jaringan reseksi otot infundibulum jantung diambil kemudian dilakukan isolasi untuk menilai viabilitas sel dan ekspresi genetik. Hasil: Delapan subjek ((n = 4) ± 95% C.I) kelompok suhu 37°C secara signifikan menghasilkan jumlah sel viabel yang lebih banyak (mean: 2675sel/mg) dibandingkan suhu 4°C (mean: 970 sel/mg) dengan (p <0,05). Ekspresi gen yang mengekspresikan sel kardiomiosit secara flowsitometer terlihat kelompok medium transpor 37°C secara bermakna lebih tinggi dibandingkan dengan suhu 4°C. Simpulan: Isolasi kardiomiosit menggunakan medium transpor suhu fisiologis (37⁰C) menghasilkan jumlah sel viable yang lebih banyak dibandingkan medium konvensional (4⁰C).

---

Background: Coronary heart disease is one of the main causes of death in developing countries, including Indonesia. This disease is one of the causes of heart failure. The therapy that has been implemented has not able to repair the damage of the heart muscles that have occurred. Stem cell therapy, either injection or patch, has not succeeded satisfactorily. The use of the cardiac patch is exposed to many problems such as low viability of the cells produced. Hypothermia at 4°C, which is used for isolation of cardiomyocytes related with activity disturbance which causes a decrease in the number of viable cells produced. The temperature 37°C which is the body's physiological temperature considered can produce better cell viability.

Methods: An experimental invitro study was conducted in the Pelayanan Jantung Terpadu Cipto Mangunkusumo (PJT-RSCM) and the Indonesian Medical Education and Research Institute (IMERI) Faculty of Medicine, University of Indonesia from December 2019 to November 2020 by recruiting patient subjects who underwent total correction of tetralogy of Fallot. The resection of the cardiac infundibulum was taken and then isolated to assess cell viability and gene expression.

Results: A total of eight subjects ((n = 4) ± 95% CI) the 37°C group produced significantly more viable cells (mean: 2675 cells/mg) than at 4°C (mean: 970 cells/mg).)) with (p <0.05). The expression of genes expressing cardiomyocytes by flowcitometer showed that the physiological group (37°C) was significantly higher than the conventional group (4°C).

Conclusion: Isolation of cardiomyocytes using a physiological temperature transport medium (37⁰C) resulted in a higher number of cells than conventional medium (4⁰C)."

"ABSTRAKPenggunaan sel punca sebagai anti fibrosis hati cukup menjanjikan. Sel punca CD34 asal darah tali pusat sudah banyak digunakan dalam studi anti fibrosis hati. Penelitian ini menjelaskan efek ko-kultur antara sel stelata hepatik HSC LX-2 dan sel punca CD34 asal darah tali pusat dalam morfologi sel dan ekspresi TGF-?, tenascin-C dan kolagen tipe 1A1. Metode : Sel CD34 diisolasi dari sel darah tali pusat manusia yang dikriopreservasi menggunakan separasi magnet. Sel HSC LX-2 dikultur sebagai kontrol monokultur. Sebagian dipanen dan dihitung untuk dilakukan ko-kultur dengan sel CD34 dalam rasio 1:1. Ko-kultur CD34 dan LX-2 dilakukan dengan metode kultur konvensional 2D dan 3D hanging drop. Hasil monokultur dan ko-kultur dipanen pada hari ke1, 2 dan 3 dan dilakukan pewarnaan imunositokimia tenascin-C ekstraksi RNA untuk analisis kuantitatif dengan real time PCR ekspresi TGF-? dan kolagen tipe 1A1.Hasil : Hasil menunjukkan perbedaan morfologi ko-kultur 2D dan 3D hanging drop dibandingkan kontrol monokultur. Pada ko-kultur 2D terdapat mikromassa, sedangkan pada monokultur 2D tidak ada mikromassa yang terbentuk. Pada ko-kultur 3D hanging drop, terdapat spheroid yang lebih kecil hambatan pembentukan spheroid dibandingkan monokultur 3D hanging drop. Sel CD34 memiliki efek direk terhadap aktivitas sinyal sel stelata hepatik dengan adanya kecenderungan penurunan ekspresi TGF-?. Analisis imunositokimia tenascin-C dalam mikromassa dan spheroid masih perlu dioptimasi. Ko-kultur 2D dan 3D hanging drop method sel punca CD34 asal darah tali pusat dan sel stelata hepatik memiliki efek terhadap penurunan ekspresi kolagen tipe 1A1.Kesimpulan : Sel punca CD34 asal darah tali pusat memiliki efek direk terhadap morfologi sel, inhibisi aktivitas sel stelata hepatik LX-2 yang ditandai dengan penurunan ekspresi TGF-beta dan inhibisi deposisi matriks ekstrasel yang ditandai penurunan ekspresi kolagen tipe 1A1.Kata kunci: sel punca asal darah tali pusat CD34 , sel stelata hepatik, liver fibrosis, TGF-beta, tenascin-C, kolagen 1A1.

---

ABSTRACTBackground The development of stem cell therapy antifibrotik placing as one of the promising therapy. Umbilical cord blood CD34 stem cells has been widely used in the study antifibrosis. This study describes the effect of co culture between hepatic stellate cells HSC LX 2 and umbilical cord blood CD34 stem cells on cell morphology and expression of TGF , tenascin C and collagen type 1A1.Method CD34 cells were isolated from thawed cryopreserved human umbilical cord blood cells using magnetic separation. LX 2 cells culture were harvested and counted. CD34 and LX 2 cells were mixed in suspension with 1 1 ratio v v . Cell suspension divided into 2 sets 2D co culture plated in standard well plate and 3D co culture as hanging drops. LX 2 monoculture, CD34 dan LX 2 coculture were harvested on day 1, 2 and 3 as sample for further analysis. Tenascin C expression was analysed by imunocytochemistry techniques. TGF Beta and collagen type 1A1 expression was analysed by qPCR.Result The result showed different morphology between co culture and monoculture on 2D and 3D hanging drop. The 2D co culture showed micromass formation, instead of no micromass formation on monoculture. The 3D hanging drop showed smaller spheroid formation spheroid formation inhibition compared with monoculture. CD34 cells showed direct effect on hepatic stellate cell signalling activity represented by the decrease in TGF beta expression, inhibition of extracellular matrix deposition represented by a decrease in Collagen type 1A1 expression.Conclusion UCB CD34 cells showed direct effect on cell morphology, inhibition of hepatic stellate cell LX 2 activity represented by a decrease in TGF beta expression, inhibition of extracellular matrix deposition represented by a decrease in collagen type 1A1 expression. "

"Diferensiasi osteogenik dari Sel punca mesenkim (MSC) menjadi osteoblas memiliki signifikansi klinis yang sangat penting untuk mengobati cedera tulang. Berbagai penelitian telah dilakukan untuk meneliti faktor-faktor yang dapat meningkatkan diferensiasi osteogenik, termasuk pengembangan perancah untuk kultur MSC. Perancah Polivinil Alkohol (PVA)/ Fibroblast-derived Matrix (hFDM) asal manusia menjadi salah satu kandidat perancah yang diduga dapat mendukung diferensiasi osteogenik MSC. Keberadaan matriks ekstraseluler (ECM) pada perancah dapat meregulasi berbagai aktivitas seluler melalui komponen protein matriks yang terdapat pada ECM. Protein matriks berperan sebagai sekuesterasi berbagai faktor pertumbuhan. Faktor pertumbuhan seperti BMP2 dan chordin diketahui dapat meregulasi diferensiasi osteogenik. Proses terjadinya diferensiasi osteogenik dapat diamati melalui akumulasi mineral kalsium yang terdeposit pada matriks ekstraseluler. Tujuan dari penelitian ini adalah untuk mengetahui metode optimum dalam pembuatan perancah PVA/hFDM, danmengetahui peran perancah PVA/hFDM dalam mempengaruhi diferensiasi osteogenik MSC dengan mengukur ekspresi gen BMP2, dan chordin, serta ekspresi kadar kalsium relatif pada matriks ekstraseluler. Optimasi pembuatan perancah PVA hFDM dimulai dengan optimasi medium kultur, waktu kultur, preparasi, dan teknik deselulerisasi. hFDM dikarakterisasi menggunakan pewarnaan Hematoxylin, Masson Trichrome, dan Imunohistokimia untuk mengetahui keberadaan protein matriks. MSC dikultur pada perancah PVA/hFDM untuk uji diferensiasi osteogenik selama 21 hari. Sampel RNA diisolasi pada hari ke-7,14, dan 21. Ekspresi gen BMP2 dan chordin dianalisis menggunakan metode qRT-PCR. Adapun ekspresi kadar kalsium relatif dianalisis dengan uji kualitatif dan kuantitatif pewarnaan Alizarin Red. Hasil penelitian ini menunjukkan protokol pembuatan perancah PVA/hFDM telah dioptimasi, dan karakterisasi hFDM memperlihatkan keberadaan protein matriks berupa kolagen dan biglycan. Ekspresi gen BMP2 menurun pada kelompok MSC yang dikultur pada perancah PVA/hFDM baik di hari ke-7, 14, dan 21. Sedangkan ekspresi gen chordin meningkat pada kelompok MSC yang dikultur pada perancah PVA/hFDM di hari ke 7, dan 14, kembali menurun di hari ke-21. Ekspresi kadar kalsium relatif cenderung meningkat pada kelompok MSC yang dikultur pada perancah PVA/hFDM dengan gambaran mikroskopis berupa bercak merah pada permukaan perancah. Kesimpulan dari penelitian ini adalah perancah PVA/hFDM cenderung dapat mendukung diferensiasi osteogenik MSC. Hasil penelitian menunjukkan bahwa penggunaan perancah PVA/hFDM dapat menurunkan ekspresi gen BMP2, dan meningkatkan ekspresi gen chordin, serta cenderung meningkatkan ekspresi kadar kalsium relatif yang terdeposit pada matriks ekstraseluler.

---

Osteogenic differentiation from Mesenchymal Stem Cell (MSC) to osteoblasts has great clinical significance for treating bone injury. Various studies have been conducted to investigate factors that can enhance osteogenic differentiation, including scaffold development for MSC culture. Scaffold Polyvinyl Alcohol (PVA) / human Fibroblast-derived Matrix (hFDM) is a scaffold candidate assumed to support osteogenic differentiation of MSCs. The extracellular matrix (ECM) presence on the scaffold can regulate various cellular activities through the matrix protein components contained in the ECM. Matrix protein plays a role in sequestering multiple growth factors. Growth factors such as BMP-2 and chordin are to regulate osteogenic differentiation. The process of osteogenic differentiation can be observed by accumulating calcium minerals in the extracellular matrix. The purpose of this study was to determine the optimal method for making PVA / hFDM scaffold and to determine the role of the PVA / hFDM scaffold in affecting MSC osteogenic differentiation by measuring the expression of BMP2 and chordin genes, as well as the expression of relative calcium levels in the extracellular matrix. Optimization of making hFDM PVA scaffold begins with the optimization of culture medium, culture time, preparation, and decellularization techniques. hFDM was characterized using Hematoxylin, Masson Trichrome, and Immunohistochemical staining to determine matrix proteins' presence. MSCs were cultured on the PVA / hFDM scaffold for osteogenic differentiation assay for 21 days. RNA samples were isolated on day 7,14 and 21. Expression of BMP2 and chordin genes were analyzed using the qRT-PCR method. The expression of relative calcium levels was analyzed by qualitative and quantitative tests of Alizarin Red staining. The results of this study indicate that the PVA / hFDM scaffold preparation protocol has been optimized, and the hFDM characterization shows the presence of matrix proteins in the form of collagen and biglycan. BMP-2 gene expression decreased in the MSC group cultured on the PVA / hFDM scaffold on days 7, 14, and 21. In contrast, the chordin gene expression increased in the MSC group cultured on the PVA / hFDM scaffold on days 7, and 14, back down on day 21. The expression of relative calcium levels tended to increase in the MSC group cultured on the PVA / hFDM scaffold with a microscopic appearance of red spots on the scaffold surface. This study concludes that Scaffold PVA / hFDM tends to support osteogenic differentiation of MSCs. The results showed that the use of the PVA / hFDM scaffold could decrease the expression of the BMP2 gene, and increase the expression of the chordin gene, and tended to increase the expression of the relative calcium levels deposited in the extracellular matrix.

"

"The main objective of this book is to provide a comprehensive review on stem cells and their role in tissue regeneration, homeostasis and therapy. In addition, the role of cancer stem cells in cancer initiation, progression and drug resistance are discussed. The cell signaling pathways and microRNA regulating stem cell self-renewal, tissue homeostasis and drug resistance are also mentioned. Overall, these reviews will provide a new understanding of the influence of stem cells in tissue regeneration, disease regulation, therapy and drug resistance in several human diseases."

"Ekstrak flavonoid yang terkandung didalam propolis telah terbukti dapat meningkatkan fungsi jantung pasca myocardial infarction (MI). Penelitian ini bertujuan untuk mempelajari pengaruh propolis terhadap karakteristik swelling, profil rilis, degradasi, dan toksisitas hidrogel polivinil alkohol (PVA)-gelatin untuk aplikasi perancah patch jantung. Perancah hidrogel PVA/gelatin difabrikasi menggunakan metode freeze thaw dengan penambahan propolis sebanyak 3%, 7%, dan 10%. Inkorporasi propolis didalam matriks hidrogel menyebabkan penurunan swelling ratio hidrogel menjadi sekitar 254%, 221% dan 190% saat penambahan propolis sebanyak 3%, 7%, dan 10% secara berurutan. Kemampuan swelling ini mampu menjadikan hidrogel sebagai sistem penghantar obat yang melepas propolis melalui mekanisme sustained released. Dalam durasi 6 jam, perancah hidrogel mampu melepas propolis sebanyak 4,30%, 4,86%, dan 5,68% seiring dengan meningkatnya kandungan propolis 3%, 7%, dan 10%.  Penambahan konsentrasi propolis terbukti memodifikasi laju degradasi hidrogel dimana seiring penambahan propolis, weight loss yang diamati semakin tinggi. Sampel dengan propolis 3%, 7%, dan 10% mengalami pengurangan berat sebanyak 31%, 41%, dan 48% secara berurutan. Degradasi yang terjadi pada hidrogel mengikuti mekanisme surface erosion sehingga memampukan patch terdegradasi dalam lingkungan biologis seiring perbaikan jaringan jantung. Hasil uji sitotoksisitas mendapati nilai viabilitas sel pada kadar propolis 3%, 7% dan 10%, adalah 77%, 94%, dan 80% secara berurutan.  Nilai viabilitas sel menunjukkan bahwa propolis tidak menghambat metabolisme sel HEK-293 dan tidak bersifat toksik. Penelitian ini menunjukkan propolis dapat dienkapsulasi ke dalam matriks hidrogel sebagai sistem penghantaran obat maupun sebagai perancah patch jantung yang berpotensi mempercepat regenerasi jaringan baru.

---

Flavonoid extracts contained in propolis have been shown to improve heart function after myocardial infarction (MI). This study aims to study the effect of propolis on swelling characteristics, release profile, degradation, and toxicity of hydrogels made from polyvinyl alcohol (PVA)-gelatin for cardiac patch scaffold applications. The PVA/gelatin hydrogel scaffolds were fabricated using the freeze thaw method with the addition of 3%, 7% and 10% propolis. The incorporation of propolis in the hydrogel matrix led to a decrease in the swelling ratio of the hydrogel to around 254%, 221% and 190% when the concentration of propolis was 3%, 7% and 10% respectively. This swelling behavior turns the hydrogel into a drug delivery system that releases propolis through a sustained release mechanism. Within 6 hours, the hydrogel scaffolds were able to release 4.30%, 4.86%, and 5.68% of propolis as the propolis concentration increased by 3%, 7%, and 10%. The addition of propolis concentration has been shown to modify the hydrogel degradation rate as when propolis is added, the observed weight loss is higher. Samples with 3%, 7%, and 10% propolis experienced a weight reduction of 31%, 41%, and 48%, respectively. The degradation that occurs in the hydrogel follows the surface erosion mechanism so that it enables the patch to degrade in a biological environment as cardiac tissue repairs. Cytotoxicity test results found cell viability values at propolis levels of 3%, 7% and 10% were 77%, 94% and 80% respectively. This research shows that propolis can be incorporated into a hydrogel matrix as a drug delivery system or as a cardiac patch scaffold which has the potential to accelerate the regeneration of new tissue."

"ABSTRAKNama :Nugraha Utama PelupessyProgram Studi :S3 Ilmu KedokteranJudul :Marker Cancer Stem Cells CD133, CD44, dan ALDH1A1 Sebagai Faktor Prognostik pada Kanker Ovarium Tipe Epitelial Kanker ovarium merupakan penyakit yang bersifat heterogen dan kebanyakan pasien datang dengan stadium lanjut. Kanker ovarium epitelial tipe II mempunyai sifat pertumbuhan tumor yang cepat dan secara genetik labil dibandingkan tipe I. Keberadaan cancer stem cells CSC dianggap sebagai salah satu faktor prognostik terjadinya kemoresisten dan kesintasan hidup yang rendah.Penelitian ini bertujuan untuk membuktikan CSC sebagai faktor prognostik dengan menggunakan marker CD133, CD44, dan ALDH1A1 pada kanker ovarium tipe epitelial.Marker CD133, CD44, dan ALDH1A1 diperiksa dengan imunohistokimia dan flowcytometry. Hasil ekspresi marker CSC pasien kanker ovarium tipe I dan tipe II dimasukkan kedalam suatu tabel yang dihubungkan dengan respons kemoterapi dan kesintasan hidup. Analisis data dilakukan dengan program computer STATA 14. Analisis kesintasan dilakukan dengan analisis Kaplan-Meier dan uji asumsi cox proportional hazard. Analisis multivariat dipakai untuk model prognosis selama 10 bulan. Sistem skoring dibuat dengan menggunakan receiver operating characteristic ROC curve analyses.Data demografi kelompok terbanyak adalah usia ge; 45 tahun; 40 sampel 72,7 , stadium I, 23 sampel 41,8 , diferensiasi buruk 30 sampel 54,5 , dan tipe II 16 sampel 29,1 . Perbedaan yang bermakna antara tipe histopatologi dengan marker CSC hanya terlihat pada marker CD44. Skor Prediksi Kemoresisten SPKr 10 bulan yang dihubungkan dengan 4 variabel yaitu usia ge; 45 tahun, tipe II, stadium III minus;IV, dan CD44 tinggi dengan ROC 72,47 dan probabilitas post test 82,5 . Kurva ROC berdasarkan kombinasi marker CSC dan faktor klinikopatologi yaitu stadium III minus;IV, usia ge; 45 tahun, diferensiasi buruk, tipe II, CD133 negatif, CD44 tinggi, dan ALDH1A1 tinggi adalah 0,841. Skor Prediksi Kematian SPKm 10 bulan yang dihubungkan dengan 3 variabel yaitu stadium III minus;IV, tipe II, dan CD44 tinggi dengan AUC 80,44 dan probabilitas post test 78,7 . Kurva ROC berdasarkan kombinasi marker CSC dan faktor klinikopatologi yaitu stadium III minus;IV, usia ge; 45 tahun, diferensiasi buruk, tipe II, CD133 positif, CD44 tinggi, dan ALDH1A1 tinggi adalah 0,841.Simpulan: Marker CD44 terbukti berperan pada kanker ovarium tipe II. Skor Prediksi Kemoresisten dan Skor Prediksi Kematian dapat ditentukan selain dengan faktor klinikopatologi, juga dengan memakai marker CSC. Kata kunci: ALDH1A1, CD44, CD133, CSC, kanker ovarium epitelial, kesintasan hidup, respons kemoterapi.

---

ABSTRACTName : Nugraha Utama PelupessyStudy Program : Doctoral Program Medical SciencesTitle :Cancer Stem Cell CD133, CD44 andALDH1A1 Markers As Prognostic Factors on Epithelial Ovarian Cancer. Ovarian cancer is a heterogeneous disease and most of the patients came with an advanced stage. Epithelial ovarian cancer type II has the characteristic of rapid tumor growth and genetically more labile than that of type I. The presence of cancer stem cells CSC is considered as one of the prognostic factors of low mortality and survival.The aims of this study was to prove CSC as prognostic factors using CD133, CD44, and ALDH1A1 markers on epithelial ovarian cancer.Clinicopathology and demographic data were collected from medical records. CD133, CD44, and ALDH1A1 markers were examined with flowcytometry and immunohistochemistry. CSC marker expression of the patients with ovarian cancer type I and II was connected with chemotherapy and survival response. Data analysis was done by using STATA 14 software. Survival analysis was done by using Kaplan-Meier analysis and Cox proportional hazard test. Multivariate analysis is used for prognosis model for ten months. Receiver Operating Characteristic ROC curve analyses was used as the system scoring. The highest group demographic data were age ge; 45 years; 40 samples 72.7 , stage I, 23 samples 41.8 , poor differentiation 30 samples 54.5 , and type II 16 samples 29.1 . A significant difference between the histopathologic type and the CSC marker was seen only in CD44 marker. Chemoresistance Prediction Score in 10 months was associated with 4 variables ie age ge; 45 years, type II, stage III minus;IV, and CD44 high with ROC 72.47 and posttest probability 82.5 . The highest chemoresitency scoring ROC curve based on the combination of CSC marker and clinicopathology factors; stage III minus;IV, age ge; 45 years, poor differentiation, type II, negative CD133, high CD44, and high ALDH1A1, was 0.841. Mortality Prediction Score in 10 months was associated with 3 variables is stage III minus;IV, type II, and CD44 high with AUC 80.44 and posttest probability 78.7 . The highest mortality scoring ROC curve based on the combination of CSC marker and clinicopathology factors; stage III minus;IV, age ge; 45 years, poor differentiation, type II, positive CD133, high CD44, and high ALDH1A1, was 0.841. Conclusion: The CD44 marker has a role in type II ovarian epithelial cancer. Chemoresistance Prediction Score and Mortality Prediction Score can be determined from clinicopathological factors and using CSC marker. Keywords: ALDH1A1, CD44, CD133, chemotherapy response, CSC, Epithelial Ovarian Cancer, survival"