ADIPOCYTE DERIVED STEM CELLS AND Mesenchymal Product as REGENERATIVE

MEDICINE

Indah Yulianto.,Endra Yustin., (Sebelas Maret University)., Titi Moertolo (Trisakti University).,Dewi Margono (Medical Dentist Indonesia University).,Yuyun R A(National Seoul University).,Yuda H Fibrianto (Medical Veterinary Gajah Mada University)

By definition, a stem cell is characterized by its ability to self-renew and its ability to differentiate along multiple lineage pathways.

Ideally, a stem cell for regenerative medicinal applications should meet the following criteria:

Regenerative Medicine :Uses the body’s own stem cells and growth factors is an alternative therapeutic strategy for repair of damage tissue, and becoming predominant cell base therapy

Can be found in abundant quantities (millions to billions of cells)

Can be harvested by a minimally invasive procedure

Can be differentiated along multiple cell lineage pathways in a regulatable and reproducible manner.

Can be safely and effectively transplanted to either an autologous or allogeneic host

Regenerative medicinal applications should meet the following criteria:

Human subcutaneous adipose tissue depots as a potential source of adult or somatic stem cells;

With the increased incidence of obesity in the Indonesia and abroad, subcutaneous adipose tissue is abundant and readily accessible.

Approximately 400,000 liposuction surgeries are performed all over the world each year,

These procedures yield anywhere from 100 mL to >3 L of lipoaspirate tissue

Hasil lemak dari liposuction

pellet

Adipocyte after 24 hours

Adipocyte after 24 hours

ADSC pasage 1 : hari ke 3

A. ADSCs hari ke 7 dan B. ADSC hari ke 14

A B

by dr Indah Yulianto – Yuyun Rindiastuti – Dewi M

Transdifferensiasi ADSCs to Odontoblast(immunocitokimia)

Ditujukan untuk regenerasi dan renewal Pulpitis

Courtesy by Indah Yulianto – Yuyun Rindiastuti – Dewi M

Preparing product Mesenchymal from Adipocyte

ADSC which are a population of pluripotent mesenchymal cells and possess the ability to differentiate into various lineages

ADSCs secrete various growth factors that repair and replace the defective surrounding cells

Also a good raw material for skin regeneration, reepithelialization, wound healing and wrinkling care instead of ADSC itself

Advance ADSCs Protein Extract by Definitions

Oxygen levels & exogenous

oxidative stress

Genetic stability

Cytogenetic analysis, gene

expression, bioenergetics

studies

Culture grown in physiologycal 20% oxygen

Culture grown in standard 3%

oxygen

-transcriptional activation in HIF 1A

target genes- Inhibition oxygen

consumption

- Increase genetic instability

- Decrease glycolysis

Oxygen sensing,

HIF 1 response

RESULTGrowth of hMSC at 20% O2 reduces lifespan, increases oxidative stress and the rate of telomere shortening

Telomerase shortening &

cell senescence

Cell growth

Growth of hMSC at 20% O2 increases DSB generation and chromosomal instability.

DSB level (53BP1 marker)

DNA damage & chromosomal

aberration

Exogenous source of ROS cause growth

defects & genetic

instability

DISCUSSION

Culture of mammalian cells at 20%

oxygen

- Promotes DNA damage & senescence

- Chromosomal aberration during long term culture

Importance of biosafety

in cell therapy

Culture ADMSC at 3% 0xygen

Increase growth rate

- No evidence of spontaneous immortalization

-- reduce telomere shortening rate decrease structural chromosomal aberration

Significant upregulation of gene expression in cells grown at 3%

oxygen tension related to HIF 1A stabilization

Balance bioenergetic

s pattern

Implication of gene ontology analysis (HIF

1 dependent gene expression)

Autocrine & paracrine effect in

cell growth & senescence

HIF 1A decreases OXPHOS system by

downregulating mitochondrial ATP

production & oxygen consumption

Analysis Method

Cell number & viability

Trypan blue stained cell counted by automatic cell

counting machine (invitrogen)

MSC marker CD 44, CD 106 PCR

Energy metabolism gene related glycolisis

mechanism

HIF 1 A, HIF 1B, GLUT-1 PCR

Pluripotency marker

HIF 2A, Oct-4, Sox-2, Rex-1

PCR

Courtesy by Yuyun Rindiastuti

RESULT

Cell proliferation & viability

syn-chro-

nization

Normoxic 24h

Hypoxic 24h

Normoxic 48h

Hypoxic 48h

Normoxic 72h

Hypoxic 72h

0

500,000

1,000,000

1,500,000

2,000,000

2,500,000

3,000,000

3,500,000

4,000,000

4,500,000

5,000,000

Graph 1. Cell number profile

Cell

num

ber

syn-chro-

nization

Normoxic 24h

Hypoxic 24h

Normoxic 48h

Hypoxic 48h

Normoxic 72h

Hypoxic 72h

86%

88%

90%

92%

94%

96%

98%

Graph 2. Cell viability

Cell

via

bili

ty

Courtesy By Yuyun RA

MSC surface marker expression

HIF 1A & GLUT 1 expression

Normoxic group Hypoxic group

Courtesy By Yuyun RA

3 days after skin needling + MCSs + product

Acne and Acne Scar 1 week After Skin Needling + MCSc + Product

1 week After Skin Needling + MCSs + Product

Acne and Acne Scar

Courtesy by Titik Moertolo

Alopecia Universalis : 1 month after MChSc and Product

Alopecia Universalis :1 month after MCSc + Product MSCc

By Indah Yulianto

After 2 Months

After :2 Months

By Indah Yulianto

After 4 months

After 4 months

By Indah Yulianto

(1)Luka iris dan (2) penyembuhan 2 hari pasca olesan krem produk

MCSc

1

2

Courtesy by dr Titik Moertolo

Acne Papulo pustular + Scar Acne : before and 1 week after AAPE

(3) Scar acne pasca skin needling + product & (4) 1 week after skin needling + MCSs +_ Product

34

Acne Papulo Pustular + Scar

Courtesy by dr Titik Moertolo

1 month after Skin Needling + MShCs

Acne Scar & Box scar 1 week post Dermaroller

Acne Scar before (Box Scar)2 weeks after Skin Needling +

MShCs

Courtesy by dr Titik Moertolo

Infant 2 weeks, with fistula supra ani

Tissue regeneration after 1 week using MshCs Product - Cream

BY Indah Yulianto

Chronical Ucler diabeticum > 3

years

Day three, after injection with

MSCHs product

1 week1 month after Mesenchymal

product ( Injections and Cream) – Men 74 years old

By Indah Yulianto

Leprosy after 30 years + 3 months burning ulcer +

Infection

3 weeks after MSCs Product

Injections + Cream

BY Indah Yulianto

Chronical Ulcer in M Hansen (Malperforan dupied) more than 20

years

1 week after

2 weeks after (painful & sensitivity +) 25 % reduce in 2 weeks

Bone

Luka bakar + infeksi 5 bulan y.l.

intra kutan produk MShCs

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

regenerasi

Suport By : Derm Ma Ma Biotechnology Laboratory