STEM SEL DAN PRODUK STEM SEL UNTUK … · Klinik Bayi Tabung “GladioolIVF” di RSIA Gladiool Magelang PT. Dermama Indonesia PT. Wersut Seguni Indonesia. The Stem Cell Concept A

STEM SEL DAN PRODUK STEM SEL UNTUK PENGOBATAN REGENERATIVE

Drh Yuda Heru Fibrianto MP PhDBagian Fisiologi FKH-UGM YogyakartaKlinik Bayi Tabung ldquoGladioolIVFrdquo di RSIA Gladiool MagelangPT Dermama IndonesiaPT Wersut Seguni Indonesia

The Stem Cell Concept

A stem cell is an undifferentiated

dividing cell that gives rise to a

daughter cell like itself and a

daughter cell that becomes a

specialized cell type

Properties of a stem cell

Must have telomerase

Has active genes that inhibit differentiation

Has active genes that promote growth and cell division

Often polarized enabling it to give rise to disperate progeny

Embryonic Stem cell Adult Stem cell

Pluripotency

Defined as the ability of a cell to differentiate into any cell type of the three germ layers

The acquisition of pluripotency in somatic cells requires the activation of a transcriptional regulatory network (this involves NANOG SOX2 OCT4 TCF3 and possibly others)

Teratoma formation is the most stringent test for pluripotency that is available in the human system

Embryonic Stem Cells

Embryonic stem cells are derived from the inner cell mass of a preimplantation blastocyst

These pluripotent stem cells can give rise to all tissues including the complete spectrum of mesoderm endoderm and ectoderm derivatives

Human ESCs were first isolated a decade ago and have now been demonstrated to give rise to various cell types including hematopoietic cells neuron-like cells glialprogenitors dendritic cells hepatocytes pancreatic islet-like cells osteocytes chondrocytes adipocytes cardiomyocytes as well as muscular endothelial skin lung and retinal tissues

This quintessential differentiation potential provides a promising avenue to produce a large quantity of transplantable cells from a renewable source

Perinatal Stem Cells There is growing evidence to support the diverse differentiation capacity of

stem cells derived from perinatal sources including umbilical cord blood (UCB)

Transplantation of UCB has been clinically successful for hematopoietic stem cell applications resulting in high degree of engraftment favorable immunotolerance and limited evidence for graft-versus-host disease compared to adult bone marrow stem cell transplantation

UCB-derived stem cells are capable of in vitro expansion long-term maintenance and differentiation into representative cells of all three embryonic germinal layers that is the endoderm (eg hepatopancreaticprecursor cells mature hepatocytes and type II alveolar pneumocytes) mesoderm (eg adipocytes chondrocytes osteoblasts myocytes and endothelial cells) and ectoderm (eg neurons astrocytes and oligodendrocytes) (Van de Ven et al 2007)

UCB may provide a clinically applicable that avoids ethical challenges raised with embryonic sources

Amniotic epithelial cells (AECs) from amniotic membranes can also be induced to differentiate into diverse and specialized cell types from all three germ layers including pancreatic cells (endoderm) cardiomyocytes(mesoderm) and keratinocytes (ectoderm)

Adult Stem Cells

Adult stem cells comprise a wide range of progenitors derived from nonembryonic non-fetal tissues such as bone marrow adipose tissue and resident stem cell pools

Adult stem cells are a leading candidate for clinical application in regenerative medicine based on accessibility autologous status and favorable proliferative potential

Bone marrow-derived stem cells are a cornerstone of contemporary regenerative medicine applications

Bone marrow-derived hematopoietic stem cells represent the earliest example of cell-based regenerative medicine pioneered to address the needs of patients treated with total body irradiation for leukemia that developed life-threatening infections and irreversible tissue destruction

Stem cells defined by expression of the CD34 surface marker can also be obtained via peripheral blood leukapheresis for clinical engraftment

Provided the foundation for autologous and allogeneic stem cell transplantation and offer novel treatments for patients with cancer autoimmune diseases and genetic diseases including severe combined immunodeficiency and thalassemia

Transplant studies have also revealed engraftment of nonhematopoietic cell lineages derived from donor bone marrow unmasking subpopulations capable of a diverse range of lineage-specific differentiation

Mesenchymal stem cells Also discovered in the bone marrow although at low frequency compared to the

hematopoietic pool isolated from connective components of various postnatal tissues including adipose and synovial tissue as well as from peripheral and cord blood

Mesenchymal stem cells exhibit properties of multipotency with the capacity to contribute to regeneration of bone cartilage and muscle tissues of mesodermalorigin (Phinney and Prockop 2007)

Evidence also supports the contribution of mesenchymal stem cells to liver and pancreatic islet cell regeneration and protection in the setting of kidney heart or lung injury

Mesenchymal stem cells secrete a spectrum of bioactive molecules that provide a regenerative microenvironment to limit the area of damage and to mount a self-regulated regenerative response

Autologous and allogeneic mesenchymal stem cells have been tested in recent clinical trials including for treatment of osteogenesis imperfecta Crohnrsquosdisease and graft-versus-host disease

Bioengineered Stem Cells Embryogenesis is a sequential process of differential gene expression

dictated by the epigenetic environment Exploiting epigenetic influence on phenotypic outcome biotechnology platforms are developed for reversal of differentiation to achieve genetic reprogramming of adult sources back to an embryonic state

Such platforms include ldquotherapeutic cloningrdquo and ldquonuclear reprogrammingrdquo that bypass the need for embryo extraction to generate pluripotent stem cell phenotypes from autologous sources

Reprogramming of adult stem cells to generate customized embryonic-like stem cells offers the future for patient-specific regenerative therapies

The following three issues are important purposes of ES and iPS cell researches for regenerative medicine 1) Dissection of differentiation mechanisms 2) Application to cell transplantation and 3) Drug discovery

Embryo-like stem cell

Embryonic stem cell from somatic cell nuclear transfer (SCNT) therapeutic cloning

Partenogenesis

iPS (induced pluripotent stem cell)

G0 stage

Oocyte Donor

Collection of cell donor

In vitro Maturation

Culture Nuclear Transfer

+_

Enucleation

In vitro CultureEmbryo development

Electrofusion

2 3 45

6

5-6 hari

7

1

89

Patient

Patient

Cloned human ES cells for therapeutic process

Embryonic stem cell

Imunological

compatible

Chemical activation

iPS production with Transcription Factor Transfection

OCT 34 C-Myc Sox 2 Nanog

Retrovirus vector

AP-staining

62 Day

Alasan pemilihan sel mesenchimal dan sel

lemak sebagai sumber sel punca

Sifat autologous (Bang et al 2005 Jung et al 2009 Kang et al 2006)

MSC mudah diperoleh dari jaringan pasien sendiri termasuk sumsum tulang jaringan adiposa tulang rawan sinovium periosteum otot dan amandel palatine dan dapat diperluas sampai berdiferensiasi menjadi garis keturunan sel tertentu

Cont

Sejumlah penelitian telah menunjukkan bahwa sumsum tulang berisi populasi sel punca mesenchymal yang mampu berdifferensiasi menjadi beberapa garis keturunan termasuk jaringan neuron dan glia (Jiang et al 2002 2003 Nagai et al 2007 Kim et al 2008)

Pengadaan sumsum tulang sangat menyakitkan bagi pasien jumlah sel yang dipanen hasilnya rendah dan terbatas oleh usia pasien

Sel punca mesenchimal asal jaringan adiposa adalah sumber alternatif yang baik untuk terapi sel karena akses mudah dan jumlah sel yang tinggi serta mempunyai respon yang baik pada kultur untuk pemulihan kemampuan proliferasi dan memperbaharui diri sendiri dibanding dengan MSC dari sunsum tulang (Efimenko et al 2011)

Jaringan lemak seperti juga sumsum tulang berasal dari mesoderm embrionik dan mengandung populasi sel stroma heterogen yang mencakup adipocytes masak preadipocytes fibroblas pembuluh darah sel-sel otot polos sel endotel monosit makrofag dan limfosit (Caspar-Bauguil et al 2005)

Sel punca yang diturunkan dari jaringan lemak (ADSCs) mempunyai kemampuan untuk memperbaharui diri sendiri dan dapat berdifferensiasi sepanjang garis keturunan beberapa jaringan mesenkim termasuk adipocytes osteoblast miosit kondrosit sel endotel dan kardiomiosit (Peng et al 2008 Lin et al 2008 Park et al 2008 Zuk et al 2002)

Dapat berdiferensiasi menjadi sel saraf fungsional menggunakan bFGF dan forskolin (Jang et al 2010)

Cont

Differentiation of HESC in vitroGrowth factors Non-protein

factors

Feeder layer

Neurons NGF EGF bFGF PDGF IGF-

1 NT-3 BDNF TGFα fgf-8

SHH

RA MS5 (stromal cell) feeder

MedII (human hepato carcinoma) condition

media

Glia EGF BFGF PDGF IGF-1

NT-3 BDNF

RA

Kidney NGF

Bone Dexamethasone

ascorbic acid β-

glyserophosphate

Cardiomyocyte TGFβ 5-aza-2rsquo-deoxycytidine END-2 (visceral endoderm) feeder layer

Endothelial cell VEGF PDGF

Blood cells Cytokine + BMP4 + VEGF C166 (yolk sac endothelia)

S17 (bone marrow) feeder layer

Liver NGF HGF aFGF Sod butyrat Primary hepatocyte condition media

Pancreas NGF Various hormone growth factor and matrix component

NGF nerve growth factor EGF epidermal aFGF acidic fibroblast bFGF basic fibroblast FGF-8 fibroblast PDGF platelet derived HGF hepatocyte IGF-1 insulin like NT-3 neurotropin 3 BDNF brain derived TGFα transforming TGFβ RA retinoic acid VEGF vascular endothelial BMP bone morphogenic protein

Stem cell homing

Schematic representation of some of the interactions between HSC and their endosteal and vascular niches

Schematic representation of some of the interactions between HSC and their endosteal and vascular niches

Mechanisms of stem cell function after homing into the damaged heart

Application and research of stem cells

Embrionic stem cell (ESC) yang menurunkan progenitor sel saraf (ESNPs) adalah satu di antara kandidat jenis sel saraf yang paling menjanjikan yang sedang diselidiki untuk memperbaiki SSP

Sel mampu mempertahankan potensi untuk berkembang biak dan berdiferensiasi menjadi subtipe neuron dan glia setelah pencangkokan (Tabal et al 2005) dengan hasil tertentu tergantung pada arahan lingkungan lokal (Carpentino et al 2008 Maisano et al 2009)

Begitu sel berdiferensiasi akan membentuk neuron fungsional yang mampu berhubungan dan menyatu ke dalam otak penerima (Ruschenschmidt et al 2005)

Neurosurgery 2011 Jan68(1)213-22 discussion 222

Predifferentiated brain-derived adult human progenitor cells migrate toward ischemia after transplantation to the adult rat brain

Olstorn H Varghese M Murrell W Moe MC Langmoen IA

The adult human brain contains neural stemprogenitor cells (AHNPCs) that can survive transplantation into the adult rat brain migrate toward a lesion and display limited neuronal differentiation in vivo

PLoS One 2011 Mar 46(3)e17560

Functional integration of grafted neural stem cell-derived dopaminergic neurons monitored by optogenetics in an in vitro Parkinson modelToslashnnesen J Parish CL Soslashrensen AT Andersson A Lundberg C Deisseroth K Arenas E Lindvall O Kokaia M

Hartman et al (2010) menggunakan model tikus dari epilepsi lobus temporal telah teramati bahwa progenitor sel saraf yang diturunkan dari sel embrionik (ESNPs) bertahan dan berdifferensiasi dalam lapisan sel granula setelah melepaskan stereotaxic ke lekukan dentate (DG) menggantikan sel-sel endogen blade atas ESNPs dicangkokkan ke DG menunjukkan migrasi luas melalui blade atas sepanjang sumbu septotemporal dari hippocampus

Epilepsia 2010 Jul51 Suppl 371-5

Effect of neuronal precursor cells derived from medial ganglionic eminence in an acute epileptic seizure model

Calcagnotto ME Ruiz LP Blanco MM Santos-Junior JG Valente MF Patti C Frussa-Filho R Santiago MF Zipancic I Alvarez-Dolado M Mello LE Longo BM

Epilepsia 2010 Jul51 Suppl 366-70

Grafting of GABAergic precursors rescues deficits in hippocampal inhibition

Calcagnotto ME Zipancic I Piquer-Gil M Mello LE Alvarez-Dolado M

Stem Cells 2010 Jul28(7)1153-64Medial ganglionic eminence-derived neural stem cell grafts ease spontaneous seizures and restore GDNF expression in a rat model of chronic temporal lobe epilepsyWaldau B Hattiangady B Kuruba R Shetty AK

ScienceDaily (Aug 30 2008) mdash Oregon Health amp Science University scientists have successfully produced functional auditory hair cells in the cochlea of the mouse inner ear The breakthrough suggests that a new therapy may be developed in the future to successfully treat hearing loss The results of this research was recently published by the journal Nature

Human embryonic stem cells modified to glow red when the stem cells become red blood cells ScienceDaily (Nov 29 2009) mdash Victorian stem cell scientists from MonashUniversity have modified a human embryonic stem cell (hESC) line to glow red when the stem cells become red blood cells

ScienceDaily (Aug 3 2009) mdash University of Florida researchers were able to program bone marrow stem cells to repair damaged retinas in mice suggesting a potential treatment for one of the most common causes of vision loss in older people

Modification of radiation-induced oral mucositis (mouse) by adult stem cell therapy single-dose irradiation Schmidt M1 Piro-Hussong A Siegemund A Gabriel P Doumlrr Win Radiat Environ Biophys 2014 Jun 15

Mol Cancer Ther 2014 Jun 13 pii molcanther01752014 Human umbilical cord blood- derived mesenchymal stem cells producing IL-15 eradicate established pancreatic tumor in syngeneic mice Jing W1 Chen Y2 Lu L3 Hu X4 Shao C4 Zhang Y4 Zhou X4 Zhou Y4 Wu L5 Liu R4 Fan K6 Jin G4

Cardiol Rev 2014 Jun 12

Cardio-Oncology Issues among Pediatric Cancer and Stem Cell Transplant Survivors Hochberg JC1 Cairo MS Friedman DM

Int J Hematol 2014 Jun 14

Combination of bortezomib thalidomide and dexamethasone (VTD) as a consolidation therapy after autologousstem cell transplantation for symptomatic multiple myeloma in Japanese patients Takashima S1 Miyamoto T Kadowaki M Ito Y Aoki T Takase K Shima TYoshimoto G Kato K Muta T Shiratsuchi M Takenaka K Iwasaki H Teshima TKamimura T Akashi K

Blood Cells Mol Dis 2014 Jun 9 pii S1079-9796(14)00054-0 doi 101016jbcmd201405003 Hematopoietic stem cells and liver regeneration Differentially acting hematopoietic stem cell mobilization agents reverse induced chronic liver injury

Tsolaki E1 Athanasiou E2 Gounari E1 Zogas N1 Siotou E2Yiangou M3 Anagnostopoulos A2Yannaki E4

HASIL REPRODUKSI

Tekhnik kedokteran tinggi

Sekretome

Faktor-faktor yang diproduksi oleh sel stem tersebut disebutsekretome mikrovesikel atau eksosome

Bahan-bahan yang digunakan untuk bertahan hidup dan tumbuh

Beberapa penelitian terhadap faktor-faktor yang disekresikanoleh sel stem memperlihatkan telah mampu memperbaikijaringanorgan yang semula dalam kondisi rusak

Fakta bahwa sel stem mensekresikan bermacam-macam faktor pertumbuhan juga telah diperlihatkan oleh para peneliti proteomik dengan membuktikan keberadaan berbagai macam faktor pertumbuhan dan sitokin lainnya dalam ekstrak media penumbuh sel stem

Platelet Derived Growth Factor (PDGF) Interleukine (IL) ndash 6 dan 15 Transforming Growth Factor (TGF) β Epidermal Growth Factor (EGF) dan Leukimia Inhibitory Factor (LIF)

After 4 months

After 2 months

After 1 months

After 4 monthsAfter 3 months

Acne Papulo Pustular + Scar

Courtesy by dr Titi Moertolo

Acne Scar amp Box scar

1 week post Dermaroller

2 weeks after Skin Needling + MShCs

Chronical Ulcer diabeticum gt 3 years

1 month after Mesenchymal product (Injections and Cream) ndash Men 74 years

old

1 week

Leprosy after 30 years + 3 months burning ulcer

+ Infection

3 weeks after MSCs Product Injections +

Cream

BY Indah Yulianto

Luka bakar + infeksi 5 bulan ylintra kutan produk MShCs

Pengolesan Krem + Produk MShCsHari ke 15 penyembuhan 75 tanpa Cicatriks ataupun Retraksi persendian

regenerasi

Fistula supra ani

Duchene muscular dysthrophysyndrome

Car accident

After 3 times

Diabetes Melitus

Osteoarthritis

DMD syndrom

Cerebral palsi