Cells: Mitochondria, Glycocalyx
vs.
Free radical processes
… a brain storming session …
with hypotheses & speculations
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Cell anatomy
Cells differ tremendously in
morphology, but the main features
of mammalian cells are quite similar
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N = from some dozens to many thousands
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Mitochondrion
Mitochondria of an adult absorb about
400 liters of oxygen per day
Mitochondria (sing. mitochondrion)
are known as the powerhouses of
the cell, producing the majority about 95% - of the ATP that is used
in cell metabolism
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Oxyhaemoglobin
Cell membrane
Cytosol
molecules of dioxygen O2
Matrix
O2 <≡> ●O–O● e→ ●O–O <≡> O2●
+
O2● e→ O–O 2H
→ HO–OH → H2O2
HO–OH <≡> HO●●OH e→ HO +
●OH
e
2H+
HO + ●OH → HO + OH → 2H2O
O2 + 4e + 4H+ → 2H2O
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Mitochondrion ROS; O2●
It has been estimated that 0.1% to 2.0% of O2
consumed by mitochondria generates O2●
There are the following leak events; corresponding to 0.4-8.0 liters O2 per day:
• Complex I leaks O2● towards the matrix
• Complex III leaks O2● towards the intermembrane space
• Complex III leaks O2● towards the matrix
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Mitochondrion ROS; H2O2
Long-living H2O2 molecules are “freely” movable
(even to cytosol)
• H2O2 can attack the components in the intermembrane space
• H2O2 can attack the components in the matrix
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mROS leak (O2● ; H2O2)
Nature = genius architect !!!
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Scavenging mROS
ROS production should be controlled, not stopped
Human = genius being !!!
mito-Antioxidant
(+)
Cytosol
mTA
Cell
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Mitochondrially targeted
antioxidant
+P
… can be substituted e.g. by +N
1x
X … is an efficient antioxidant
MitoSNO1
SkQ2M
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Chemical structure of
SkQ1
a promising compound
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MitoQ vs. SkQ1
Selective targeting of a redox-active ubiquinone to mitochondria within cells: antioxidant and antiapoptotic properties.
J.Biol.Chem. 276 (2001) 4588-4596
2e ; 2H+
MitoQuinone
MitoQuinol
2e ; 2H+
Plastoquinone
SkQ1
SkQ1 is based on plastoquinone, an antioxidant from plant chloroplasts (2004)
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mTA series
N+
P+
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V. P. Skulachevs’ offer
The main purpose of the Institute of Mitoengineering is to organize largescale research projects aimed at development and practical application of
novel highly effective drugs
Maxim Vladimirovich
Skulachev; biologist
Director of the Institute of Mitoengineering
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Skulachev’s tenets
1. The “ideal” antioxidant should be specifically targeted to
mitochondria where ROS are produced.
2. This antioxidant should effectively remove not all the ROS
but just their excess.
3. It is also important for an antioxidant not to be toxic and not
to be recognized and eliminated by cell enzymes.
4. Unfortunately none of the antioxidants discovered by the end of
the 20th century fits all these criteria.
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Skulachev’s observations
1.
Extremely high effectiveness of SkQ1 (hundreds-fold higher when
compared to analogic MitoQ) can be explained by SkQ1 ability to “get
regenerated”.
2.
Having accomplished its function as ROS neutralizer and changed into an
oxygenated form, SkQ1 is easily restored by a mitochondrial respiratory
chain.
3.
It is due to this ability for multiple restorations that SkQ1 proves to be
effective even in extremely low nanomolar concentrations.
4.
SkQ-type compounds allows them to act as the recyclable antioxidants, in
contrast to conventional antioxidants that are usually inactivated after
interaction with a free radical.
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L. Šoltés idea
Constructing & patenting the “Tamer’s membranes”
SkQ1
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Patent pending - confidential
+
mTA
Biomembrane + mTA
Wound healing
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Preclinical test for wound dressing
containing antioxidant
Healing time
3 days
7 days
10 days
14 days
18 days
21 days
Control
Complex
hydrogel
Hydrogel
+
antioxidant
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Cells: Mitochondria, Glycocalyx
vs.
Free radical processes
… a brain storming session …
with hypotheses & speculations
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… a tissue …
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… detail of the cell surface …
Extracellular fluid
nm
Cytosol
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The outer cell space
The protein and lipid cell membrane
is covered with a layer of
carbohydrate chains
The content of carbohydrate chains
is about 3%
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Glycocalyx
Glycocalyx is a sticky cell
envelope composed of oligoand poly-saccharides that
surround the cells
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Glycocalyx functions

Has a negative electrical charge which repels
negatively charged ions

Protect cells against damages

Serve to attach cells to one another

Act as a part of the receptor sites

Enter into immune reactions
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Glycocalyx functions

Has a negative electrical charge which repels
negatively charged ions

Protect cells against damages

Serve to attach cells to one another

Act as a part of the receptor sites

Enter into immune reactions
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“Glycocalyx” protective function
?
Fibroblasts in synovium
HAs of MDa molar mass move into the synovial fluid and protect cartilage from damages
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“Glycocalyx” protective function
!!!
HAs of MDa molar mass
ECM with HAs of MDa molar mass protect cartilage chondrocytes against damages
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Glycocalyx functions

Has a negative electrical charge which repels
negatively charged ions

Protect cells against damages

Serve to attach cells to one another

Act as a part of the receptor sites

Enter into immune reactions
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“Glycocalyx” in cell attachment
!!!
HMM
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Glycocalyx functions

Has a negative electrical charge which repels
negatively charged ions

Protect cells against damages

Serve to attach cells to one another

Act as a part of the receptor sites

Enter into immune reactions
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“Glycocalyx” vs. immune reactions
!!!
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“Glycocalyx” - masking tumor cells
by native HMM HAs !!!
immunogenic
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“Glycocalyx” - masking tumor cells
Native HMM HAs ………..….
= not immunogenic
“Unmasking” tumor(s): Lower-molar-mass HAs …..
stimulates immunity responses

Depolymerizing action of HYALs ???

Degrading action of ROS/RNS !!! … papers of Šoltés et al.
Liberation of HMM HAs by lower-molar-mass hyaluronans,
which are nontoxic and not immunogenic … Šoltés speculation
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“Glycocalyx” - unmasking tumor cells
Lower MM HAs ……
= immunogenic
One of the current trends
Lower-molar-mass HAs …..
stimulates immunity responses
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.
Generation of OH
biogenic components
… degrading action of ROS …
H
H
O
O
HO-HC
+
HO-H2C
Cu(II) + O2
[100]
O
HO-HC
O
HO-H2C
[1]
H
O
O
H
O
O
+
e-
Cu(I)
O
O
O
O
AscH-
H
O
H
O
HO-HC
+ Cu(II)
HO-H2C
+ H2O2
O
O
+H+
HO-HC
- e-
HO-H2C
O
O
Cu(I)
O
O
H
DHA
H2O2 + Cu(I)---complex
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OH + Cu(II)catalyst
40
Hyaluronan degradation
step 1
CH3OC
OH
C
HO
O
O
OC
O
HO
OH
C
NH
C
H OC
HO
OH
+ HO
OH
C
HO
O
O
OC
OH
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O
HO
C
NH
C
O
O
O
O
CH3OC
OH
OH
H
CH3OC
.
OC
C
O
HO
NH
OH
OH
OH
H
O
HO
O
O
O
HO
O
NH
OH
CH3OC
OH
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Hyaluronan degradation
step 2
CH3OC
OH
C
HO
O
O
OC
O
HO
OH
C
NH
C
OH
OH
H
OC
O
C
O
HO
O
O
HO
O
NH
OH
CH3OC
OH
+ O2
OH
C
HO
O
O
OC
OH
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CH3OC
O
HO
C
NH
C
O
H
.O
OH
OH
O OC
C
O
HO
O
O
HO
O
NH
OH
CH3OC
OH
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Hyaluronan degradation
step 3
CH3OC
OH
C
HO
O
O
OC
O
HO
C
O
O
OH
O
HO
O
HO
O
NH
OH
CH3OC
CH3OC
O
HO
C
NH
C O
O
+
OH
OH
HO
O OC
C
HO
O
O
HO
O
NH
OH
CH3OC
OH
fragment 3.1
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O
C
OH
OH
OC
C
O
OH
HO
O OC
NH
C
OH
OH
HO
fragment 3.2
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VEGA project
2011-2014
 Degrading
action of ROS/RNS on HMM HAs
papers of Šoltés et al.
OH
C
HO
O
O
OC
OH
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CH3OC
O
HO
C
OH
OH
NH
C
O
OC
O
HO
O
O
HO
O
NH
OH
CH3OC
OH
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Publishing activity
4 years
Papers;
CC & WOS rating
Chapters
in books (USA)
16
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Acknowledgements
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