X-Message-Number: 32564
Date: Sun, 18 Apr 2010 13:36:40 -0700 (PDT)
From:
Subject: First Evidence That Chitosan Could Repair Spinal Damage
[Membrane damage is a component of vitrification solution toxicity. Could
chitosan nanoparticles completely block this?]
First Evidence That Chitosan Could Repair Spinal Damage
ScienceDaily (Apr. 16, 2010) - Richard Borgens and his colleagues from the
Center for Paralysis Research at the Purdue School of Veterinary Medicine have a
strong record of inventing therapies for treating nerve damage. From Ampyra,
which improves walking in multiple sclerosis patients to a spinal cord simulator
for spinal injury victims, Borgens has had a hand in developing therapies that
directly impact patients and their quality of life.
Another therapy that is currently undergoing testing is the use of polyethylene
glycol (PEG) to seal and repair damaged spinal cord nerve cells. By repairing
the damaged membranes of nerve cells, Borgens and his team can restore the
spinal cord's ability to transmit signals to the brain.
However, there is one possible clinical drawback: PEG's breakdown products are
potentially toxic. Is there a biodegradable non-toxic compound that is equally
effective at targeting and repairing damaged nerve membranes? Borgens teamed up
with physiologist Riyi Shi and chemist Youngnam Cho, who pointed out that some
sugars are capable of targeting damaged membranes. Could they find a sugar that
restored spinal cord activity as effectively as PEG? Borgens and his team
publish their discovery that chitosan can repair damaged nerve cell membranes in
the April 16 issue of The Journal of Experimental Biology.
Having initially tested mannose and found that it did not repair spinal cord
nerve membranes, Cho decided to test a modified form of chitin, one of the most
common sugars that is found in crustacean shells. Converting chitin into
chitosan, Cho isolated a segment of guinea pig spinal cord, compressed a
section, applied the modified chitin and then added a fluorescent dye that could
only enter the cells through damaged membranes. If the chitosan repaired the
crushed membranes then the spinal cord tissue would be unstained, but if the
chitosan had failed, the spinal cord neurons would be flooded with the
fluorescent dye. Viewing a section of the spinal cord under the microscope, Cho
was amazed to see that the spinal cord was completely dark. None of the dye had
entered the nerve cells. Chitosan had repaired the damaged cell membranes.
Next Cho tested whether a dose of chitosan could prevent large molecules from
leaking from damaged spinal cord cells. Testing for the presence of the colossal
enzyme lactate dehydrogenase (LDH), Borgens admits he was amazed to see that
levels of LDH leakage from chitosan treated spinal cord were lower than from
undamaged spinal cords. Not only had the sugar repaired membranes at the
compression site but also at other sites where the cell membranes were broken
due to handling. And when the duo tested for the presence of harmful reactive
oxygen species (ROS), released when ATP generating mitochondria are damaged,
they found that ROS levels also fell after applying chitosan to the damaged
tissue: chitosan probably repairs mitochondrial membranes as well as the nerve
cell membranes.
But could chitosan restore the spinal cord's ability to transmit electrical
signals to the brain through a damaged region? Measuring the brain's response to
nerve signals generated in a guinea pig's hind leg, the duo saw that the
signals were unable to reach the brain through a damaged spinal cord. However,
30.min after injecting chitosan into the rodents, the signals miraculously
returned to the animals' brains. Chitosan was able to repair the damaged spinal
cord so that it could carry signals from the animal's body to its brain.
Borgens is extremely excited by this discovery that chitosan is able to locate
and repair damaged spinal cord tissue and is even more enthusiastic by the
prospect that nanoparticles of chitosan could also target delivery of
neuroprotective drugs directly to the site of injury 'giving us a dual bang for
our buck,' says Borgens.
Story Source:
Adapted from materials provided by Journal of Experimental Biology, via
EurekAlert!, a service of AAAS. Original article written by Kathryn Knight.
Journal Reference:
First published online April 16, 2010
Journal of Experimental Biology 213, 1513-1520 (2010)
Published by The Company of Biologists 2010
doi: 10.1242/jeb.035162 This Article
Chitosan produces potent neuroprotection and physiological recovery following
traumatic spinal cord injury
Youngnam Cho1,*, Riyi Shi1,2 and Richard B. Borgens1,2
1 Center for Paralysis Research, School of Veterinary Medicine, Purdue
University, West Lafayette, IN 47907, USA
2 Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN
47907, USA
Accepted 10 January 2010
Chitosan, a non-toxic biodegradable polycationic polymer with low
immunogenicity, has been extensively investigated in various biomedical
applications. In this work, chitosan has been demonstrated to seal compromised
nerve cell membranes thus serving as a potent neuroprotector following acute
spinal cord trauma. Topical application of chitosan after complete transection
or compression of the guinea pig spinal cord facilitated sealing of neuronal
membranes in ex vivo tests, and restored the conduction of nerve impulses
through the length of spinal cords in vivo, using somatosensory evoked potential
recordings. Moreover, chitosan preferentially targeted damaged tissues, served
as a suppressor of reactive oxygen species (free radical) generation, and the
resultant lipid peroxidation of membranes, as shown in ex vivo spinal cord
samples. These findings suggest a novel medical approach to reduce the
catastrophic loss of behavior after acute spinal cord and brain injury.
Biomed Pharmacother. 2010 Feb 26. [Epub ahead of print]
Potential role of N-Succinyl-Chitosan in immune reconstitution after umbilical
cord blood transplantation in mice.
Luo H, Li J, Chen X. Department of Pharmacology, Gansu College of Traditional
Chinese Medicine, No 35, Dingxi East road, Lanzhou 730000, PR China.
Abstract
How to shorten the immune convalescence required after transplantation in
recipients is still an austere medicinal problem. Speed-up recovery means
less infection opportunity and high survival. In this study, an
immune-deficient mouse model was developed by lethally irradiation to
evaluate the immune reconstitution effect of N-Succinyl-Chitosan (NSC), a
water soluble low molecular weight chitosan derivative, after umbilical cord
blood stem cells transplantation, including hematopoiesis parameters
detection, T-cell phenotype analysis and pathological comparison; effects of
NSC on proliferation of umbilical cord blood stem cell has also been
evaluated in vitro. By comparison and analysis, we found the combination of
NSC and hematopoietic growth factors could dramatically shorten the time
required for the recovery of hematopoiesis, T-cell phenotype and injured
spleen after transplantation, suggesting that NSC may hasten the immune
recovery and therefore may shorten the time of exposure to life-threatening
opportunistic infections in transplant recipients. Moreover, we demonstrated
that NSC was effective on the proliferation and clone of umbilical cord
stem cell in vitro, suggesting NSC could speed up the immune reconstitution
in umbilical cord blood transplantation by enlarge the number of umbilical
cord blood stem cell. Copyright C 2010 Elsevier Masson SAS. All rights
reserved.
PMID: 20378302
J Biol Eng. 2010 Jan 29;4(1):2.
Chitosan nanoparticle-based neuronal membrane sealing and neuroprotection
following acrolein-induced cell injury.
Cho Y, Shi R, Ben Borgens R. Center for Paralysis Research, School of Veterinary
Medicine, Purdue University, West Lafayette, IN 47907, USA.
Abstract
ABSTRACT: BACKGROUND: The highly reactive aldehyde acrolein is a very potent
endogenous toxin with a long half-life. Acrolein is produced within cells
after insult, and is a central player in slow and progressive "secondary
injury" cascades. Indeed, acrolein-biomolecule complexes formed by
cross-linking with proteins and DNA are associated with a number of
pathologies, especially central nervous system (CNS) trauma and
neurodegenerative diseases. Hydralazine is capable of inhibiting or reducing
acrolein-induced damage. However, since hydralazine's principle activity is
to reduce blood pressure as a common anti-hypertension drug, the possible
problems encountered when applied to hypotensive trauma victims have led us
to explore alternative approaches. This study aims to evaluate such an
alternative - a chitosan nanoparticle-based therapeutic system. RESULTS:
Hydralazine-loaded chitosan nanoparticles were prepared using different
types of polyanions and characterized for particle size, morphology, zeta
potential value, and the efficiency of hydralazine entrapment and release.
Hydralazine-loaded chitosan nanoparticles ranged in size from 300 nm to 350
nm in diameter, and with a tunable, or adjustable, surface charge.
CONCLUSIONS: We evaluated the utility of chitosan nanoparticles with an
in-vitro model of acrolein-mediated cell injury using PC -12 cells. The
particles effectively, and statistically, reduced damage to membrane
integrity, secondary oxidative stress, and lipid peroxidation. This study
suggests that a chitosan nanoparticle-based therapy to interfere with
"secondary" injury may be possible.
PMID: 20205817 [PubMed - in process]PMCID: PMC2824642
Free Text>
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2824642/pdf/1754-1611-4-2.pdf
[Chitosan nanoparticles appear to be quite safe.]
Cornea. 2010 Mar 23. [Epub ahead of print]
Ocular Tolerance to a Topical Formulation of Hyaluronic Acid and Chitosan-Based
Nanoparticles.
Contreras-Ruiz L, de la Fuente M, Garcia-Vazquez C, Saez V, Seijo B, Alonso MJ,
Calonge M, Diebold Y.
>From the *Ocular Surface Group-IOBA, University of Valladolid, Valladolid,
Spain; daggerCIBER de Bioingenieria, Biomateriales y Nanomedicina (CIBER-BBN),
Valladolid, Spain; and double daggerNANOBIOFAR Group, Department of
Pharmaceutical Technology, University of Santiago de Compostela, Santiago de
Compostela, Spain.
Abstract
PURPOSE:: Hyaluronic acid-chitosan nanoparticles (HA-CS NPs) have the
potential to serve as a reliable drug delivery system to topically treat
ocular surface disorders. We evaluated the in vivo uptake by ocular
structures, the acute tolerance, and possible alterations of tear film
physiology in rabbits. METHODS:: Fluorescent HA-CS NPs (fl-HA-CS NPs) were
prepared by ionotropic gelation using fluoresceinamine-labeled hyaluronic
acid and resuspended in buffer. fl-HA-CS NPs (30 muL, 0.5 mg/mL) and
fluoresceinamine-HA conjugate (30 muL) were instilled into rabbit eyes every
30 minutes for 6 hours. In vivo uptake and acute tolerance were
characterized 24 hours after the first instillation and compared with
preinstillation measurements. Clinical signs, including tear production,
lacrimal drainage system patency and reflux, and ocular surface pathology,
were evaluated. RESULTS:: The rabbits showed no signs of ocular discomfort
or irritation after exposure to HA-CS NPs. No macroscopic alteration in
ocular surface structures was observed. fl-HA-CS NPs were present inside
conjunctival and corneal epithelial cells, although the distribution within
the cells was different. The fl-HA-CS NPs had no significant effects on
tissue morphology and functionality, tear production, or drainage.
CONCLUSION:: Taken together, these data demonstrate that HA-CS NPs are a
safe drug carrier for ocular surface application.
PMID: 20335805
[With the right nanoparticle, transport of chitosan into the brain is feasible.]
Biomaterials. 2010 Feb;31(5):908-15. Epub 2009 Oct 22.
Trimethylated chitosan-conjugated PLGA nanoparticles for the delivery of drugs
to the brain.
Wang ZH, Wang ZY, Sun CS, Wang CY, Jiang TY, Wang SL. Department of
Pharmaceutics, Shenyang Pharmaceutical University, Liaoning, China.
Abstract
Trimethylated chitosan (TMC) surface-modified poly(d,l-lactide-co-glycolide)
(PLGA) nanoparticles (TMC/PLGA-NP) were synthesized as a drug carrier for
brain delivery. TMC was covalently coupled to the surface of PLGA
nanoparticles (PLGA-NP) via a carbodiimide-mediated link. The zeta potential
of TMC/PLGA-NP was about 20mV with a mean diameter around 150nm. 6-coumarin
loaded PLGA-NP and TMC/PLGA-NP were injected into the caudal vein of mice,
and fluorescent microscopy of brain sections showed a higher accumulation of
TMC/PLGA-NP in the cortex, paracoele, the third ventricle and choroid
plexus epithelium, while no brain uptake of PLGA-NP was observed. There was
no pronounced difference in cell viability between TMC/PLGA-NP and PLGA-NP
as shown by MTT assay. Behavioral testing showed that the injection of
coenzyme Q(10) loaded TMC/PLGA-NP greatly improved memory impairment,
restoring it to a normal level, but the efficacy was slight for loaded
PLGA-NP, without TMC conjugation. The senile plaque and biochemical
parameter tests confirmed the brain-targeted effects of TMC/PLGA-NP. These
experiments show that TMC surface-modified nanoparticles are able to cross
the blood-brain barrier and appear to be a promising brain drug delivery
carrier with low toxicity.
PMID: 19853292
[Disruption of cell membranes by DMSO can occur at low concentrations. I wonder
what the effect of chitosan nanoparticles would be on hemolysis?]
PDA J Pharm Sci Technol. 2001 Jan-Feb;55(1):16-23.
Comparative hemolytic activity of undiluted organic water-miscible solvents for
intravenous and intra-arterial injection.
Mottu F, Stelling MJ, Rufenacht DA, Doelker E. School of Pharmacy, University of
Geneva, Switzerland.
Abstract
In humans, nonaqueous solvents are administered intravascularly in two kinds
of situations. They have been used in subcutaneous or intramuscular
pharmaceutical formulations to dissolve water-insoluble drugs. The need for
these vehicles had increased in recent years, since the drug development
process has yielded many poorly water-soluble drugs. The use of
water-miscible nonaqueous solvents in therefore one of the approaches for
administering these products as reference solutions useful in formulation
bioequivalence studies. The intravascular use of organic solvents has also
gained importance owing to a new approach for the treatment of cerebral
malformations using precipitating polymers dissolved in water-miscible
organic solvents. At present, the solvent most commonly used for the liquid
embolics to solubilize the polymers is dimethyl sulfoxide, which exhibits
some local and hemodynamic toxicities. In order to find new, less toxic
vehicles for pharmaceutical formulations for the intravenous and
intra-arterial routes and for embolic materials, 13 water-miscible organic
solvents currently used (diluted with water) for pharmaceutical
applications, were evaluated in this study. Their hemolytic activity and the
morphological changes induced when mixed with blood (1:99, 5:95, 10:90
solvent:blood) were estimated in vitro. From these data, the selected
organic solvents could be subdivided into four groups depending on their
hemolytic activity: very highly hemolytic solvents (ethyl lactate, dimethyl
sulfoxide), highly hemolytic solvents (polyethylene glycol 200, acetone),
moderately hemolytic solvents (tetrahydrofurfuryl alcohol,
N-methyl-2-pyrrolidone, glycerol formal, ethanol, Solketal, glycofurol) and
solvents with low hemolytic activity (propylene glycol, dimethyl isosorbide,
diglyme).
PMID: 11212416
[Could chitosan nanoparticles have prevented this unexplained failure of VS4?]
Snip>"VS4 resulted in cryopreservation damage despite the fact that
cryoprotectant toxicity was low"
Cryobiology. 2007 Feb;54(1):1-12. Epub 2006 Dec 12.
Cryopreservation of rat precision-cut liver and kidney slices by rapid freezing
and vitrification.
de Graaf IA, Draaisma AL, Schoeman O, Fahy GM, Groothuis GM, Koster HJ.
Pharmacokinetics and Drug Delivery, Groningen University Institute for Drug
Exploration, A. Deusinglaan 1, 9713 AV, Groningen, The Netherlands. I.A.M.
Abstract
Precision-cut tissue slices of both hepatic and extra-hepatic origin are
extensively used as an in vitro model to predict in vivo drug metabolism and
toxicity. Cryopreservation would greatly facilitate their use. In the
present study, we aimed to improve (1) rapid freezing and warming (200
degrees C/min) using 18% Me(2)SO as cryoprotectant and (2) vitrification
with high molarity mixtures of cryoprotectants, VM3 and VS4, as methods to
cryopreserve precision-cut rat liver and kidney slices. Viability after
cryopreservation and subsequent 3-4h of incubation at 37 degrees C was
determined by measuring ATP content and by microscopical evaluation of
histological integrity. Confirming earlier studies, viability of rat liver
slices was maintained at high levels by rapid freezing and thawing with 18%
Me(2)SO. However, vitrification of liver slices with VS4 resulted in
cryopreservation damage despite the fact that cryoprotectant toxicity was
low, no ice was formed during cooling and devitrification was prevented.
Viability of liver slices was not improved by using VM3 for vitrification.
Kidney slices were found not to survive cryopreservation by rapid freezing.
In contrast, viability of renal medullary slices was almost completely
maintained after vitrification with VS4, however vitrification of renal
cortex slices with VS4 was not successful, partly due to cryoprotectant
toxicity. Both kidney cortex and medullary slices were vitrified
successfully with VM3 (maintaining viability at 50-80% of fresh slice
levels), using an optimised pre-incubation protocol and cooling and warming
rates that prevented both visible ice-formation and cracking of the formed
glass. In conclusion, vitrification is a promising approach to cryopreserve
precision-cut (kidney) slices.
PMID: 17166492
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