:: Anti-Inflammatory
Coatings for Biomaterials
NIH R01 EB000823
Principal Investigator: John A.
Frangos, Ph.D.
Agency: NIH
The prolonged inflammatory response to an implant is one of the
primary causes for the failure to integrate into tissue. The two
sources of inflammation common to almost all implants are the
foreign body response and the relative movement of the implant with
the surrounding tissue. Based on evidence in the literature and from
our research team, the inflammatory response is mediated by the
reactive oxygen species generated by macrophages, leukocytes, and
the surrounding connective tissue. Based on our findings, it is
evident that titanium dioxide and similar ceramics, even when
present as surface coatings of polymeric biomaterials, have the
ability to breakdown reactive oxygen species that have been
identified as mediators of the inflammatory response. The goal of
this Program is to develop applications for our catalytic
antioxidant ceramic technology in the biomaterials and medical
device industry. This Program, led by UCSD, consists of five
projects with ten academic and industrial partners. Project 1 will
investigate the basic mechanisms of action of metal oxides in the
catalytic breakdown of reactive oxygen species. By understanding the
fundamental reaction kinetics of the catalytic action of titanium
dioxide, catalysts of greater efficiency may be discovered. Project
2 will fabricate and characterize materials for the other four
projects. This project involves partners from Lawrence Livermore
National Labs, Drexel Univ., and UCSD. Project 3 will test the in
vivo inflammatory and foreign body response in two in vivo models; a
standard rat model and the hamster window model. This project
provides a core service to the other projects, but also investigates
fundamental mechanisms of the inflammatory response to biomaterials.
Project 4 will determine if the catalytic antioxidant ceramic
technology is able to mitigate implant-tissue strain-induced
inflammation. It will also investigate basic mechanisms of
strain-induced inflammation. Project 5 is the interface with the
medical device industry. Four Industrial Partners have been chosen
to develop to apply the technology to four different biomaterial
needs: Biosensor membranes for inplantable glucose sensors (GlySens)
and biodegradable polymers for tissue engineering (Advanced Tissue
Sciences) with reduced foreign body response; wound dressing
material with anti-inflammatory properties (3M); and. dental
materials with improved soft tissue integration (Nobel BioCare). By
the first year, we will have elucidated the catalytic mechanisms of
action of TiO,, established the ceramic coating technologies,
characterized the inflammation response in the hamster model, and
fabricated and tested dental coatings. Our overall objective is to
provide the proof-of-principle to our industrial partners, which
will encourage them to participate in more specific product
development in the second phase (years 6-10) of the BRP.
::
Shear Stress Activation of
Endothelial Membrane Function
NIH R01 HL040696
Principal Investigator: John A.
Frangos, Ph.D.
Agency: NIH
PECAM-1 confers the ability of the endothelium to sense temporal
gradients in shear stress and strain rate. To test this overall
hypothesis, we are taking an entirely integrative approach, using
molecular biology, cell biology and biochemistry, vascular and
exercise physiology. The first specific aim is to investigate the
critical sites involved in PECAM-1's interaction with G.q and eNOS
by means of molecular manipulation. The second specific aim will
investigate the role of PECAM in the mechanochemical transduction of
shear stress, temporal gradients in shear stress, strain and strain
rate in endothelial cells from PECAM knockout and wild type mice.
The third specific aim will investigate the flow and myogenic
responses in isolated arterioles from PECAM knockout and wild type
mice. The fourth specific aim will study the role of PECAM-1 in the
vascular adaptation to exercise. This investigation will test the
comprehensive hypothesis concerning the role of PECAM-1, Gaq and
eNOS in mechanochemical transduction of clinically important
hemodynamic forces in endothelial cells. If successful it will
provide the mechanistic basis on how endothelial cells sense flow in
both normal physiology and in vascular disease.
:: Interstitial Fluid Flow in Bone
Remodeling
NIH R01 AR046797
Principal Investigator: John A.
Frangos, Ph.D.
Agency: NIH/NAIMS
It has been hypothesized that fluid shear stress, induced by the
flow of interstitial fluid, mediates the response of bone to loading
and mediates modeling/remodeling. In vitro studies have demonstrated
that bone cells are stimulated by fluid shear stress, and respond
with the release of nitric oxide (NO) and prostaglandins. The
in-vivo relevance of interstitial fluid flow (IFF), however, has yet
to be established of the proposed research is to characterize three
models of altered IFF in the absence of mechanical strain, and
determine the role of nitric oxide and prostaglandins in IFF induced
bone modeling/remodeling. Specifically, (1) we will characterize the
effects of altered IFF induced by femoral vein ligation on
histomorphometry, femoral dimensions, mechanical properties, mineral
content, and mineral density in hindlimb suspended mice and rats;
(2) we will determine the role of NO and prostaglandins if IFF-mediated
modeling/remodeling by using genetically engineered mice lacking
nitric oxide synthase 2 (NOS2), nitric oxide synthase 3 (NOS3), and
cyclooxygenase 2 (COX2); (3) We will develop an externally applied
cuff to alter IFF in bone as the first step in the clinical
application of the findings. We will seek to optimize the regime of
cuff pressure application and the duration of treatment to increase
bone; (4) to further validate that IFF is altered in our rat models,
direct measurements of IFF by magnetic resonance imaging will be
performed. The long-term goal is the development of non-
pharmacological methods to counter osteopenia of disuse.
:: Mechanosensitivity of Cell
Membranes: Role of Lipid-Protein Interactions
NIH R01
HL086943
Principal
Investigator: Mirianas
Chachisvilis, Ph.D.
Agency:
NIH
Hemodynamic shear stress stimulates
number of intracellular events that both regulate vessel structure
and also influence development of vascular pathologies. The precise
molecular mechanisms by which endothelial cells transduce this
mechanical stimulus into intracellular biochemical response have not
been established yet. The central hypothesis is that the plasma
membrane of endothelial cell acts as a mechanosensitive element;
i.e. changes in physical properties of the membrane under mechanical
stress can regulate activity of membrane proteins coupled to
intracellular signaling pathways. To test this hypothesis, we will
use an integrative approach that combines time-resolved fluorescence
microscopy, biochemistry, cell biology, and membrane micromechanics.
Our preliminary experiments show for the first time that (1) when
exposed to mechanical forces, membrane lateral fluidity and
hydration levels change and (2) that increases in membrane tension
lead to activation of bradykinin G protein coupled receptor (GPCR).
The proposed research addresses the following questions: (1) which
physical properties of the lipid bilayer change in response to
mechanical perturbation, (2) which of these changes has a clear link
to function of membrane-associated proteins such as GPCRs,
G-proteins and endothelial nitric oxide synthase (eNOS), and can
mediate mechanochemical signal transduction, and (3) what are the
specific mechanisms leading to mechanically induced activation of
GPCR receptors, eNOS and G-proteins by shear stress. We will use
state-of-the-art picosecond time-resolved fluorescence, single
molecule and fluorescence correlation spectroscopy techniques to
investigate in detail what happens to the physical properties of the
lipid bilayer membrane at the molecular level under mechanical
stress and how these changes are coupled to mechanochemical signal
transduction via direct activation of the membrane associated
proteins such as GPCR's and modulation of signal amplification
cascades through G- proteins. Specifically we propose that
mechanically-induced changes in certain membrane properties such as
thickness, lateral fluidity, polarity, membrane free volume and/or
trans-membrane lateral force profile are able to initiate and
regulate conformational changes responsible for experimentally
observed response of GPCR and G protein signal transduction pathways
and eNOS activation. If successful it will provide the mechanistic
basis on how endothelial cells sense flow in both normal physiology
and in vascular disease.
:: Mechanosensory properties in the
partially obstructed guinea pig small
intestine
NIH R01
DK072616
Principal
Investigator: Hans Gregersen,
Ph.D.
Agency:
NIH
The general objective of this
proposal is to understand the biomechanical and mechanosensory
properties of the small intestine as they relate to partial small
intestinal obstruction. Guinea pigs with obstruction will be
compared to age- and sex-matched normal guinea pigs to analyze
possible differences in afferent nerve signal transduction (Specific
Aim 3) and how it relates to the geometric (Specific Aim 1) and
biomechanical remodeling (Specific Aim 2) during obstruction. The
goals are to obtain detailed morphometric and mechanical data
including muscle contraction characteristics on the normal and
obstructed guinea-pig small intestine and to relate them to the
tissue remodeling elicited by the intestinal obstruction. Hence, new
distention protocols, new theory and methods need to be used and
further developed. The abovementioned data together with afferent
nerve signals will be used to characterize the stimulus-response
function for mechanoreceptor responses. The main hypothesis is that
partial intestinal obstruction will lead to changes in the
mechanosensory properties and that a mathematical relationship can
be established between the geometric and biomechanical remodeling
and the changes in afferent nerve signal transduction. Although
previous studies have shown morphological and biomechanical
remodeling including muscle cell hypertrophy, increased stiffness of
the intestinal wall and altered motility, much remains to be
determined from a bioengineering point of view. The studies will
focus on determination of multidimensional stress-strain properties
of the small intestinal wall with the zero-stress state taking into
account, and on the integration of the passive mechanical
properties, the muscle contractile properties, and afferent nerve
signals. The studies will shed light on the relation between the
mechanical stimulus (stress and strain) and the afferent nerve
response, i.e. information on mechanoreceptor behavior will be
obtained. The rationale is that a bioengineering model is needed in
order to deal with the complexity of intestinal obstruction. A major
goal is to demonstrate that the understanding of Gl diseases can be
greatly enhanced by using a multidisciplinary approach including
material and mathematical sciences, imaging analysis, cell and
tissue bioengineering, biology, and medical science and diagnostics.
:: Nitric
Oxide protects against microcirculatory complications of malaria
NIH R01
HL087290
Principal
Investigator: Leonardo Jose De
Moura Carvalho, Ph.D.
Agency:
NIH
We propose that low nitric oxide
(NO) bioavailability mediates the microcirculatory complications of
severe malaria; NO quenching by cell-free hemoglobin (Hb) released
as an unavoidable consequence of parasite replication and low NO
production due to hypoargininemia lead to low NO bioavailability.
Vascular leak, petechial hemorrhaging, and hypotension are well
recognized complications of experimental cerebral malaria (ECM), and
the proposed studies will determine whether poor tissue oxygenation
also functions in malaria pathogenesis by altering blood flow or
functional capillary density. Our observations that (i) free
hemoglobin (Hb) is markedly elevated during ECM, (ii) free Hb
scavenges nitric oxide (NO) and (iii) marked hypoargininemia occurs
during ECM indicate that, in contrast to sepsis, malaria shock is
caused by low NO bioavailability. A major controversy in
microcirculation research is the role of NO in mediating vascular
leak and pathogenesis, and our proposed studies will define its role
in vascular leak during ECM. A key prediction of our hypothesis is
that exogenous NO should protect against ECM pathogenesis; indeed,
NO donor administration significantly (P=0.003) protects animals
from the development of disease. The markedly protected NO
donor-treated mice abrogated the vascular leak, petechial
hemorrhage, hypotension, and impaired NO mediated signaling (cGMP
levels) that were detected in saline-injected controls with ECM.
These studies will be extended to define whether NO donor
administration protects against other microcirculatory dysfunction
during ECM, such as low tissue perfusion and oxygenation (aim 1).
Adhesion of parasitized erythrocytes (pRBCs), platelets, and
leukocytes occur during ECM and deficiency of selected cell adhesion
molecules protects against malaria pathogenesis. We will interrelate
the results of the microcirculatory complications of ECM to cell
adhesion and eCAM expression to define the cellular and molecular
mechanisms whereby cell adhesion contributes to disruption of the
blood brain barrier and pathogenesis and identify whether and how
exogenous NO protects against ECM cell adhesion (aim 2). The final
aim will assess by bioassay (arteriolar dilation, and venular leak)
and actual measurement (NO electrode) whether NO bioavailability is
impaired during ECM and restored by the protective NO donor. The
response of eNOS to ECM and NO donor treatment will also be
elucidated; a detailed understanding of in vivo eNOS responses to
free Hb or to low NO bioavailability that occurs during other
diseases (sickle cell anemia) is currently lacking. Besides
providing new information about the microcirculation, the proposed
studies may lead to adjunct therapy for malaria that rescues
millions of children from death or impaired cognition. These studies
will also address long standing controversies about malaria
pathogenesis, such as whether pRBC adhesion leads to hypoxia and
multi-organ failure (sequestration hypothesis).
::
Raman Flow Cytometry for Diagnostics and Drug Discovery
NIH
R01 EB003824
Principal Investigator: John Nolan, Ph.D.
Agency: NIH
The ability to make quantitative, high throughput molecular
measurements of biological systems is a critical need for many areas
of biomedical research. This Bioengineering Research Partnership (BRP)
aims to develop a powerful new analytical platform for high
throughput screening and selection based on Raman Flow Cytometry.
This Partnership will develop new analytical instrumentation,
optically encoded polymer resins for chemical synthesis and
screening, and nanostructured materials with unique optically
properties for sensitive reporting and encoding. The new technology
will perform Raman spectroscopy on single particles in flow to
enable new applications in sensitive multiplexed detection, drug
discovery, and diagnostics. The Raman Flow Cytometry
instrumentation, and applications will be developed by a Partnership
involving engineers, biologists, and chemists from academia,
government and industry. In the first year of the Partnership, we
will modify a commercial particle sorter to detect individual Raman
vibrational bands from single particles and sort these particles
based on their optical signature. In Years 2-5, we will develop the
ability to collect and analyze complete Raman spectra from single
particles. In parallel, the Partnership will develop new encoding
and reporting strategies for multiplexed molecular analysis and
separation. This Raman Flow Cytometry technology will be applied to
the development of therapeutics and diagnostics for bacterial
pathogens and their toxins. Raman Flow Cytometry will be an
important and general new analytical and separation capability that
will impact many areas of basic and applied biomedical research in
addition to the applications proposed here.
::
Plasma Hyperviscosity for Cardiovascular Collapse
NIH R01 HL076182
Principal Investigator: Amy Tsai, Ph.D.
Agency: NIH
The long-term objective of this project is to demonstrate that
hypervious fluids are efficacious in the treatment and improved
survival from traumatic hemorrhagic shock. It is proposed to develop
a treatment for hypovolemic cardiovascular collapse based on the
infusion of high viscosity plasma expanders, which provide a novel
small-volume resuscitation that recovers microvascular perfusion for
extended periods until surgical control of bleeding is possible. The
central hypothesis is that in conditions of hypotension, and
cardiovascular collapse, high viscosity plasma restores moderate
levels of mean arterial blood pressure needed to ensure open
capillaries and tissue perfusion. Our data shows that open
capillaries are critical to tissue survival, and viscogenic plasma
expanders with tailored oncotic pressure properties restore
microvascular function and rescue the organism from hypovolemic
cardiovascular collapse. In the case of uncontrolled bleeding, these
solutions provide limited-volume resuscitation with maximum
microvascular perfusion and a gradual increase in blood pressure
thereby minimizing re-bleeding, leading to important savings of
blood transfusions, providing a new approach for dealing with
conditions in which reduced tissue perfusion jeopardizes tissue
survival in field conditions. In this project, a microcirculatory
assessment in the hamster window preparation will be used with
sophisticated and state of the art measurements of macro and
microhemodynaimcs, including local pO2 levels, capillary pressure,
and nitric oxide release. The properties of a transfusion fluid in
terms of viscosity and oncotic properties which best recovers
cardiovascular collapse will be identified in a lethal uncontrolled
bleeding model.
::
The Role of Dipole Potential in Mechanosensing
NSF
MCB 0721396
Principal
Investigator: Mirianas
Chachisvilis, Ph.D.
Agency:
NSF
The concept underlying this Small
Grant for Exploratory Research is that intrinsic electrical
properties (the dipole moment) of the lipid bilayer of a membrane
act as a mechanosensor. The hypothesis is that the dipole potential
at the lipid-water interface of a membrane can detect and change in
response to external forces generated by mechanical stress and fluid
movement. The change in dipole potential is detected by sensory
proteins embedded in the membrane and converted to chemical signals
that generate a signal cascade that is transmitted throughout the
cell. This research has the potential to irrevocably alter the view
that membrane lipids perform only a structural and protective
function.
The broader impacts of the research are the potential to change the
view of lipid bilayer properties and function, to promote
interdisciplinary research and to train students in chemo physical
techniques. A strong education and outreach program will involve
students from the University of California and High Tech High, both
in San Diego. Moreover the results of the research will be
disseminated through professional journals, at meetings, and to the
public through a WEB site.
::
Improving transcapillary transport
by reducing interstitial fluid pressure
NIH 1R03 EB006746
Principal Investigator: Ugur
Ozerdem
Agency: NIH
We have recently developed a minimally invasive, biosensor-based,
diagnostic surgical procedure for measuring interstitial fluid
pressure (IFP) in cancer. We propose to apply this new diagnostic
technique to elucidate the role of contractile pericytes in IFP and
transcapillary transport of nanoparticles in tumors. Similar to
isometric contraction of skeletal muscle, neovascular pericytes
generate contractile forces not only on the vessel walls but also on
interstitial fluid entrapped within extracellular fibers, referred
to as tissue gel. We will test whether interstitial fluid pressure
in cancer can be reduced by interfering with pericytes to improve
the convection of nanoparticle based anti-cancer drugs. We will test
whether inhibition of pericytes results in a decrease in
interstitial fluid pressure due to decreasing compressive
contractile forces elicited by pericytes. We will test whether
pericyte-NG2 proteoglycan inhibition lowers IFP and improves
convection from the plasma to the interstitial space. We will
quantify transcapillary transport by using nanoparticles as tracers.
We anticipate finding a higher transcapillary convection of
nanoparticles (simulating high molecular weight anti-cancer drugs)
and lower interstitial fluid pressure in breast tumors when
pericytes are inhibited. By combining skills and disciplines in
bioengineering, clinical physiology and microvascular sciences this
project will transform our biosensor-based diagnostic procedure into
a tangible diagnostic tool for cancer patients. The use of
ultraminiature transducer-tipped catheters as cancer interstitial
fluid pressure biosensors is innovative in light of novel use of
biosensors portfolio of National Institute of Biomedical Imaging and
Bioengineering. The role of compressive forces generated by
pericytes within breast cancer stroma has never been investigated;
which makes this proposal innovative in shedding light on the
etiology of interstitial hypertension, a significant clinical
problem in breast cancer therapy in terms of drug delivery.
::
Does nicotine exposure inhibit convective drug delivery?
TRDRP 16IT-0212A
Principal Investigator: Ugur
Ozerdem
Agency: TRDRP -Tobacco Related Disease Research Program
We have recently developed a minimally invasive, biosensor-based,
diagnostic surgical procedure for measuring interstitial fluid
pressure (IFP) in cancer. We propose to apply this new diagnostic
technique to elucidate the role of contractile pericytes in IFP and
transcapillary transport of nanoparticles in tumors. Similar to
isometric contraction of skeletal muscle, neovascular pericytes
generate contractile forces not only on the vessel walls but also on
interstitial fluid entrapped within extracellular fibers, referred
to as tissue gel. We will test whether interstitial fluid pressure
in cancer can be reduced by interfering with pericytes to improve
the convection of nanoparticle based anti-cancer drugs. We will test
whether inhibition of pericytes results in a decrease in
interstitial fluid pressure due to decreasing compressive
contractile forces elicited by pericytes. We will test whether
pericyte-NG2 proteoglycan inhibition lowers IFP and improves
convection from the plasma to the interstitial space. We will
quantify transcapillary transport by using nanoparticles as tracers.
We anticipate finding a higher transcapillary convection of
nanoparticles (simulating high molecular weight anti-cancer drugs)
and lower interstitial fluid pressure in breast tumors when
pericytes are inhibited. By combining skills and disciplines in
bioengineering, clinical physiology and microvascular sciences this
project will transform our biosensor-based diagnostic procedure into
a tangible diagnostic tool for cancer patients. The use of
ultraminiature transducer-tipped catheters as cancer interstitial
fluid pressure biosensors is innovative in light of novel use of
biosensors portfolio of National Institute of Biomedical Imaging and
Bioengineering. The role of compressive forces generated by
pericytes within breast cancer stroma has never been investigated;
which makes this proposal innovative in shedding light on the
etiology of interstitial hypertension, a significant clinical
problem in breast cancer therapy in terms of drug delivery.
