161 |
Determinants of HIV Testing in East African Communities in TorontoJohns, Ashley January 2006 (has links)
<strong>Background. </strong> Previous evidence suggests that persons who have emigrated from HIV-endemic countries experience higher rates of HIV infection and delayed diagnosis. Despite this evidence, limited research has examined HIV testing in these populations. <br /><br /> <strong>Objectives. </strong> To examine factors associated with HIV testing, as well as motivations underlying testing behaviour, within five East African communities in Toronto. <br /><br /> <strong>Methods. </strong> Secondary data analyses were conducted using cross-sectional data collected in face-to-face interviews with people from Toronto's Ethiopian, Kenyan, Somali, Tanzanian, and Ugandan communities. Logistic regression techniques were employed to assess factors associated with "ever vs. never testing," "repeat vs. non-repeat testing," and "independent vs. directive testing. " Reasons provided for testing and not testing were described. <br /><br /> <strong>Results. </strong> Individuals from all five communities were interviewed (n=270). Males were slightly over-represented (55. 9%). The average age was 35. 7 yrs (range 17-71). Three-quarters (75. 6%) of the sample had been tested for HIV. Two-thirds (65. 7%) of testers had tested more than once and 40. 7% had independently decided to get their most recent test. 71. 1% of testers reporting previous testing for immigration purposes. Testing behaviour varied greatly across communities. Ethnicity was predictive of "ever" and "repeat" testing. Risk behaviour (including multiple sex partners, concurrent sex partners, condom non-use, and/or improper condom use) was overwhelmingly not associated with testing. Fear of exposure through sexual activity was the most frequent reason for independent testing. Immigration authorities were the most common person to initiate directive testing, followed by physicians. Low perceived risk was the most common reason for not testing. <br /><br /> <strong>Conclusions. </strong> Testing rates within this population were quite high and the immigration process heavily impacted upon testing behaviour. Many determinants and motivations of testing have been identified and should be used to inform the design of interventions to promote testing behaviour in these communities. Nevertheless, many gaps have been identified by the current research and should be addressed by future research.
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162 |
Investigation of immune quiescence: assessing the role of regulatory T cells and their link with IRF-1 in HIV-exposed sero-negative individualsAbdullahi, Abdirahman 05 January 2017 (has links)
Recent research of a cohort of HIV exposed sero-negative (HESN) female commercial sex workers in Nairobi has revealed an Immune Quiescent phenotype; characterized by reduced T cell activation and higher regulatory T cells (Tregs) in peripheral blood. HESN women also express lower levels of interferon regulatory factor-1 (IRF-1), a critical regulator known to negatively impact Treg development in mice. In this study, we analyzed the functional capacity of Tregs by an in vitro depletion assay and measured functionality by flow cytometry. Data showed Tregs suppressed CD4+ and CD8+ proliferation responses. We characterized the link between Tregs and IRF-1 in HESN and observed an inverse correlation between IRF-1 expression and Treg proportions. We also established reduced expression of IRF-1 in Tregs of healthy donors by flow cytometry. In a separate study, flow cytometric analysis of high-risk sex-workers revealed that CTLA-4 expression in memory CD4+cells, not Treg frequency, was associated with HIV seroconversion. / February 2017
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163 |
Defining C3-V4 neutralisation epitopes on human immunodeficiency virus type-1 subtype c envelope glycoproteinsWibmer, Constantinos Kurt 17 January 2012 (has links)
The rational design of an HIV-1 vaccine immunogen able to induce potent, cross-reactive, neutralising antibodies remains one of the single greatest challenges in the field of vaccine research today. Roughly a dozen broadly neutralising monoclonal antibodies have been isolated to date, and their epitopes represent important vaccination targets. Interestingly, apart from three that identify over-lapping epitopes in gp41, all of the broadly neutralising monoclonal antibodies target epitopes apparent on different conformations of gp120 (including the epitopes of PG9/PG16). Thus the gp120 monomer remains the most ideal template for immunogen design. Recently, epitopes in the C3-V4 region of gp120 have been shown to be major targets for early strain-specific neutralising antibodies in subtype C infected individuals. Autologous neutralising antibodies identify vulnerable sites on the envelope, and understanding the nature of antigenic “hotspots” on gp120 will help to guide rational vaccine design. This study sought to confirm in four individuals that the C3-V4 epitope was in fact apparent on monomeric gp120, and thereafter to better characterise the nature of viral escape from these antibodies. Using magnetic beads coated with one of 16 different recombinant gp120 proteins it was confirmed that the C3-V4 response was aimed at a monomer-specific epitope in all four cases. In two instances these antibodies were shown to contribute to autologous neutralisation, while in a third the existence of quaternary structure specific antibodies that could not be adsorbed with monomeric gp120 made this link impossible. In the forth instance transfer of the C3-V4 region was shown to expose a normally occluded epitope in the CD4 binding site. This research also provided evidence for other epitopes for autologous neutralising antibodies in C3, overlapping with the CD4 binding site and V5. Lastly, by introducing relevant escape mutations into the parental recombinant gp120s and then comparing the ability of these proteins to adsorb out anti-C3 antibodies, it was shown that while these mutations conferred complete resistance to neutralisation they did not prevent the antibodies from binding to their respective epitopes. The extensive characterisation of C3-related epitopes such as those described in this research should no doubt contribute to the rational design of a gp120 based vaccine immunogen aimed at eliciting broad and potent neutralising antibody responses.
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164 |
Analysis of HIV-induced cardiomyopathy using anti-gp120 aptamersRangel Lopes de Campos, Walter 02 February 2011 (has links)
PhD, Virology, Faculty of Health Sciences, University of the Witwatersrand / HIV-associated cardiomyopathy is a multifactorial disease with a broad spectrum of aetiologies that arise due to chronic immunosupression during HIV infection. The intricate relationship between HIV infection and cardiac co-morbidity was investigated with the aid of HIV-neutralizing aptamers. These synthetic nucleic acid ligands with antibody-like properties are molecular tools with multifunctional applications ranging from drug discovery to diagnostics and therapeutics. The advent of the HIV/AIDS pandemic has naturally married the field of HIV therapy and diagnostics with that of aptamer technology. By employing a HIV-1 neutralizing aptamer, named UCLA1, raised against the viral surface envelope glycoprotein 120, I dissected some of the pathways leading to cardiomyocyte apoptosis in a cell culture system. In chapter one I investigated the potential cytotoxic effects of UCLA1 by comparing it against a panel of 17 antiretrovirals (ARVs) in clinical use with the goal of establishing a safety portfolio geared towards its use as a therapeutic agent. Using cultured human cardiomyocytes and primary peripheral blood mononuclear cells (PBMCs), I selected some of the major biological markers of ARV-induced cytotoxicity and found no measurable deleterious effect, especially when compared to other ARVs used in the same study. In chapter two, the permissiveness of cardiomyocytes to HIV infection as well as the relationship between virus-host interaction and caspase-mediated apoptosis were investigated. Non-productive, receptor and tropism-independent infection was observed, which was arrested after the reverse transcription stage. However, interaction between the virus gp120 and the host’s CXCR-4 chemokine receptor preferentially activated caspase-9 triggering robust mitochondria-mediated apoptosis. A shift from mitochondrial-initiated, caspase-9 mediated to Fas-ligand initiated, caspase-8 mediated was observed when CM were co-cultured with HIV-infected MDM. UCLA1 protected against caspase-9 mediated
vii
apoptosis but not caspase-8 mediated. Finally in chapter three I provided answers for the shift in caspase activation by showing that supraphysiological levels of IL-1β and IL-6 during HIV infection of MDM augment the effects of tumor necrosis factor (TNF). These observation provide new insight into the complex pathophysiology of HIVCM and highlight the potential of UCLA1 as a novel therapeutic agent to fight HIV and some of its associated diseases.
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165 |
The management of HIV positive patients using a CD8/38 flow cytometry assay as an alternative to viral load testingMoodley, Keshendree 10 October 2011 (has links)
MSc (Med), Faculty of Health Sciences, University of the Witwatersrand, 2011 / BACKGROUND: Human Immunodeficiency Virus (HIV) is a global epidemic with growing
numbers of people on highly active anti‐retroviral therapy (HAART) programmes.
Effectiveness of treatment needs to be monitored to ensure the uncompromised well
being of patients. This is currently done using both Viral Load (VL) and CD4 cell counts
for HAART initiation and follow‐up. Although VL is the best predictor of disease
progression it is often too expensive for monitoring patients in resource‐limited settings.
There is thus a need for a cheaper, more accessible alternative to monitor long term
patient response to therapy.
METHODS: This study evaluated the use of a recently described flow cytometric assay of
CD38 expression (previously developed at the Johannesburg Flow Cytometry Reference
Laboratory) in a cohort of HIV+ patients failing 1st line therapy, who were subsequently
enrolled onto 2nd line HAART. CD38 and CD8 were “piggy ‐backed” onto the PLG/CD4
protocol and mean fluorescence intensity (MFI) of the CD8/38 expression was
monitored longitudinally. Patterns of CD38 expression were compared to 1st line
treatment observations to establish equivalence in the predictive power of CD38
expression of fluctuation in viral load on 2nd line treatment patients. In addition, the
effect of sample age on assay accuracy was tested before implementation of the CD38
assay at a secondary testing site.
RESULTS: The patterns observed in the cohort of 2nd line therapy patients mirrored
patterns previously seen in 1st line therapy with 55% of patients showing a continuous
decline in CD38 MFI that mimicked changes in VL. The remaining 33% of patients had
non‐specific increases in CD38 MFI without concurrent increases in VL and one patient
showed irregular VL and CD38 MFI (non‐responder). The CD38 assay showed acceptable
accuracy and reproducibility up to 48 hours after venesection (%CV<5%).
Implementation at the secondary testing site was successful with 98% similarity
(%CV<5%) compared to the reference laboratory.
CONCLUSION: CD38 monitoring of 2nd line therapy patients showed comparable
patterns to observations in 1st line therapy patients. The assay proved stable over time
and easy to implement at another PLG/CD4 testing facility. As such, the CD38 assay
offers a cost‐effective, reliable real time supplementary test to long‐term VL monitoring
of HIV infected patients on the national ART programme.
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166 |
Generation of soluble, catalytically active covalent HIV-1 subtype C integrase-DNA complexes to identify novel strand transfer inhibitorsBeyleveld, Grant James January 2012 (has links)
Dissertation submitted to the Faculty of Health Sciences, University of the
Witwatersrand, Johannesburg, in fulfillment of the requirements for the degree of
Master of Science in Medicine.
Johannesburg, 2011 / The HIV-1 integrase (IN) enzyme is an integral part of the viral replication cycle
and has no known human homologues, making it an ideal target for antiretroviral
therapy. To date, only one inhibitor of IN strand transfer activity (Raltegravir,
IsentressTM) is available for human use. However, the inevitable emergence of
antiretroviral drug resistance requires ongoing research into new/novel therapies.
There are currently no assays to screen for IN inhibitors against HIV-1 subtype C
in South Africa (and worldwide), therefore, the overall objective of this study was to
generate and characterize locally relevant, soluble, functional recombinant HIV-1
subtype C IN proteins for use in strand transfer assays. Recombinant integrase
genes, including a soluble HIV-1 subtype C mutant (05ZAFV6 with C56S, C65S,
W131D, F185D and C280S) and HIV-1 subtype C Y143C mutant (05ZAFV6
soluble with Y143C) were designed, generated and cloned in frame into pET15b.
Optimal bacterial expression conditions for the expression of these constructs as
well as an HIV-1 subtype C wild type (05ZAFV6), subtype B wild type (NL4-3), and
subtype B soluble (NL4-3 with F185K and C280S; as controls) IN, in E.coli BL21
cells were determined. All five recombinant IN were successfully purified using
nickel affinity chromatography, and subsequently used to establish a strand
transfer assay to assess their activity and their response to two well-known
integrase inhibitors, L-Chicoric acid and Raltegravir. All five recombinant IN
proteins were found to be biologically active, with INY143C (116.67%) showing
equivalent activity to INBwt (117.37%), while INCsol (52.96%) was the lowest. The
IC50 values of L-Chicoric acid were higher than the expected values for all five
recombinant IN, with the subtype B and C IN solubility mutations contributing to an
increased resistance to inhibition by L-Chicoric acid.
The dose responses to Raltegravir for INCwt and INBsol were as expected, with
IC50’s in line with published data, and the INY143C mutant (known mutation
conferring resistance to Raltegravir) was resistant to inhibition of strand transfer
activity at all Raltegravir concentrations tested except the highest (50 μM).
Finally, methods to complex the INY143C mutant to thiolated-DNA were evaluated,
however definitive data could not be obtained. Future work should focus on
optimization of the purification and characterization of the IN-DNA complexes.
Overall, this study has led to the establishment of functional strand transfer assays
based on HIV-1 subtype C recombinant IN proteins, and established a framework
for screening of novel HIV-1 subtype C IN inhibitors.
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167 |
The development of a screening tool to evaluate infants who are HIV positiveHilburn, Nicole Clare 06 April 2011 (has links)
PhD, Faculty of Health Sciences, University of the Witwatersrand / HIV/AIDS continues to be one of the greatest health challenges which South Africa faces.
The epidemic in children is closely linked to that in women, the prevalence of which
continues to grow according to antenatal statistics from the South African Department of
Health (DOH). HIV is known to invade the central nervous system at the time of infection,
and causes widespead damage. In children, this leads to a well-researched condition
known as HIV encephalopathy, which affects all areas of neurodevelopment. The effects
of timely initiation of antiretroviral therapy on alleviating the impact of encephalopathy
have been well described.
Neurodevelopmental delay is a stage four disease indicator according to the World Health
Organisation (WHO), and therefore is a criterion for initiation of Highly Active Antiretroviral
Therapy (HAART). HAART is often only administered according to the virologic and
immunologic status of a child, as standardised neurodevelopmental assessment tools are
not widely available in South African clinics. When HAART initation is dependent on
immunologic status, it is often too late to prevent encephalopahy. To date, the only means
of prevention of this condition is early initation of HAART, which has not been widely
available in South Africa. Stringent guidelines for the commencement of this therapy
according to the WHO, and the South African Department of Health (DOH) have had to
be followed, leading to late initiation of HAART, and widespread central nervous system
encephalopathy. Studies which have been carried out in South African clinics have
demonstrated the high prevalence of this condition. Once there is evidence of
encephalopathy, children should be referred for assessments in all facets of development,
and where necessary, for rehabilitation. A standardised developmental screening tool
which is suitable for use in a developing country is therefore necessary in order to screen
for neurodevelopmental delays to allow for further assessment and referral to
rehabilitation services, as well as providing an additional assessment criterion for initiation
of HAART.
Paediatric HIV clinics in developing countries are understaffed, and children may be seen
by junior staff or screened by nurses due to the high numbers of clinic attendees. This
often results in neurodevelopment being inadequately assessed and children are
therefore not referred for intervention services. A standardised screening tool, which
The Development of a Screening Tool to Evaluate Infants who are HIV Positive
could be administered by clinic staff in order to ensure correct and timely referral of
children for further assessment and intervention is therefore necessary. This is of
importance both locally and internationally where a screening tool, which has been
developed specifically for this purpose, does not exist.
The aim of this study was therefore to evaluate the agreement between the Bayley-III
Screening Test and the Bayley Scales of Infant Development (3rd version) in a population
of HIV positive infants in order to evaluate its appropriateness for use in South Africa. The
Bayley Scales of Infant Development have long been considered the ‘gold standard’ in
infant developmental assessment, which is why this tool was chosen to evaluate the
Bayley-III Screening Test against. The developmental scores in each facet (cognitive,
motor or language) were evaluated to determine which should be included in an
assessment tool for this population. Further objectives for the study were to adapt the
screening tool to the needs of the population, or to develop a new screening tool should
the Bayley-III Screening Test not prove suitable for use in this population.
In order to meet the aims and objectives, a cross-sectional study was conducted where
112 HIV positive infants between the ages of six and eighteen months were assessed
using the Bayley-III Screening Test and the Bayley Scales of Infant Development (3rd
version) (BSID III). The infants were stratified into four age groups namely 6-8 months, 9-
12 months, 13-16 months, and 17-18 months. Children were recruited from Harriet Shezi
Children’s Clinic at Chris Hani Baragwanath Hospital in Soweto.
The agreement between the Bayley-III Screening Test and the Bayley Scales of Infant
Development (3rd version) was analysed using Kappa, for the overall group, and for each
age group. Overall agreement between the tools was as follows: K=0.58 for the Cognitive
facet, K=0.82 for the Expressive Communication facet, K=0.76 for the Receptive
Communication facet, K=0.44 for the Fine Motor facet and K=0.57 for the Gross Motor
facet. These values indicate that the Bayley-III Screening Test is therefore not
acceptable for clinical use, as excellent agreement (k≥0.75) in all facets would be
necessary for this purpose.
A new screening tool therefore had to be developed. The infant’s developmental scores
from the BSID III were analysed to determine which facets of development were most
severely affected, and therefore which facets should be included in a new screening tool.
Gross motor function was demonstrated to be the area which was most severely affected,
followed by cognitive function. A gross motor screening tool would therefore be suitable
for use in this population, as no equipment would be necessary. Gross motor
development is the most universally similar aspect of development, which is not
completely dependent on cultural or socioeconomic factors which often have an influence
on language and cognitive development.
Item selection from the BSID III was undertaken to determine which items should be
included in a brief screening tool. In each of the four age groups, item selection occurred
as follows: Two items which discriminated the At-Risk, from Emerging and Competent
groups (less than 20% in the At-Risk group, and 100% in the other groups) were selected.
Two items, which discriminated between children in the ‘Emerging’ and ‘Competent’
categories on the BSID III were selected (0-5% of children who were At-Risk obtained
credit, 30-50% of the Emerging group obtained credit, and 100% of the Competent group
obtained credit). Lastly, two items were selected which discriminated the Competent
group from the other two groups (100% or as high as possible in the Competent group,
and 0% in the other groups).
The new gross motor screening tool was assembled using the selected items, scoring
was allocated, and it was tested against the scores obtained on the Gross Motor facet of
the BSID III for the initial 112 infants. Agreement between the tools was analysed using
Kappa, and refinements were made according to the discrepancies. This was done three
times, until the Kappa value revealed excellent agreement between the tools (k = 0.87). A
panel of experts was then invited to examine the new gross motor screening tool, and to
comment on it, and further adjustments were made accordingly.
Preliminary concurrent validity testing of the new gross motor screening tool was then
carried out against the Gross Motor facet of the BSID III on 60 children, who were
recruited from the Harriet Shezi Children’s Clinic at Chris Hani Baragwanath Hospital in
Soweto. Statistical analysis revealed that the agreement between the BSID III and the
new screening tool was excellent (k=0.85). The diagnostic properties of the new gross
motor screening tool were as follows: sensitivity 97.4%, specificity 85.7%, positive
predictive value 92.7%, and negative predictive value 94.7%. These values indicate that the statistical properties of the tool are excellent, and the tool will not be predisposed to
underreferrals or over-referrals. Preliminary reliability testing was carried out on 15
children for test-retest/intrarater reliability and 15 children for interrater reliability.
Interrater, test-retest and intrarater reliability were excellent (r=1, r=0.98, r=0.98
respectively). Further testing of reliablity and validity should be undertaken in order to
establish these properties, and standardisation should also be carried out on healthy
children. Given the need for an assessment tool of this nature in South Africa and other
developing countries, and the statistical properties thus far, the tool may be used clinically
for the purposes for which it was developed.
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168 |
Inhibition of Human Immunodeficiency virus replication through small RNA-induced gene silencing of HIV-1 Tat specific factor 1Green, Victoria Andress 14 February 2012 (has links)
Ph.D., Faculty of Health Sciences, University of the Witwatersrand, 2011 / The
HIV-‐1
pandemic
continues
unabated.
Although
treatments
exist
that
can
substantially
alleviate
the
morbidity
and
mortality
associated
with
HIV,
there
is
still
a
need
for
improved
anti-‐HIV
treatments
that
reduce
toxicities
and
administration
frequency
and
mediate
sustained
inhibition
of
viral
replication.
Given
the
high
mutability
and
variability
of
the
virus,
a
strategy
that
is
garnering
increasing
focus
is
the
targeting
of
host
factors
that
the
virus
requires
to
replicate,
so-‐called
HIV-‐dependency
factors
(HDFs).
It
is
hoped
this
will
reduce
the
emergence
of
viral
drug
resistance.
A
number
of
genome-‐wide
screens
have
been
performed
to
identify
HDFs,
although
many
remain
to
be
validated,
particularly
in
relevant
cells
lines.
An
objective
of
this
thesis
was
to
validate
three
host
factors
as
HDFs,
in
both
TZM-‐bl
reporter
and
T
cell-‐derived
cell
lines,
and
to
examine
their
potential
as
anti-‐HIV-‐1
therapeutic
targets
through
exploitation
of
the
cellular
gene
silencing
pathway,
RNA
interference
(RNAi).
These
were
HIV-‐1
Tat
specific
factor
1
(HTATSF1),
DEAD
(Asp-‐Glu-‐Ala-‐Asp)
box
polypeptide
3,
X-‐
linked
(DDX3X)
and
SWI/SNF
related,
matrix
associated,
actin
dependent
regulator
of
chromatin,
subfamily
b,
member
1
(SMARCB1),
selected
because
they
had
been
previously
implicated
in
HIV-‐
1
pathogenesis.
The
well-‐characterised
HDF,
PC4
and
SFRS1
interacting
protein
1
(PSIP1)/lens
epithelium-‐derived
growth
factor
(LEDGF)/p75,
was
included
in
the
study
as
a
positive
control.
Cassettes
expressing
short
hairpin
RNAs
(shRNAs)
targeting
the
four
host
proteins
were
generated,
although
shRNAs
did
not
suppress
endogenous
ddx3x
mRNA
levels.
The
ability
of
shRNAs
to
inhibit
HIV-‐1
replication
in
the
reporter
cell
line,
TZM-‐bl,
was
examined.
These
HeLa-‐
derived
cells
are
permissive
for
R5-‐tropic
HIV-‐1
infection
and
contain
an
integrated
luciferase
gene
driven
by
the
viral
promoter.
shRNAs
mediated
a
dose-‐dependent
inhibition
of
luciferase
activity
in
cells
infected
with
a
HIV-‐1
subtype
B
molecular
clone
and,
although
production
of
the
viral
protein
p24
was
unaltered,
infectious
particle
production
was
decreased
in
cells
treated
with
a
shRNA
suppressing
HTATSF1.
Little
effect
was
observed
with
a
shRNA
targeting
SMARCB1,
suggesting
that
this
may
not
function
as
an
HDF
under
these
conditions.
No
effect
on
infectious
particle
production
was
seen
with
the
shRNA
targeting
PSIP1,
which
was
a
result
of
the
long
half-‐
life
of
this
protein,
highlighting
a
limitation
of
using
such
reporter
systems
for
HDF
validation.
Importantly,
shRNAs
were
not
associated
with
any
cytotoxic
effects
in
TZM-‐bl
cells.
Whether
HTATSF1
is
a
potential
therapeutic
target
was
interrogated
further
in
the
more
relevant
T
cell-‐derived
SupT1
cell
line.
Lentiviruses
were
used
to
generate
populations
where
>90%
had
one
copy
of
the
integrated
shRNA
expression
cassette.
Replication
of
the
subtype
B
molecular
clone
p81A-‐4
was
significantly
inhibited
in
the
shH1-‐expressing
SupT1
cell
line,
which
targets
HTATSF1,
for
over
14
days
post-‐infection,
although
inhibition
was
not
as
pronounced
asthat
observed
in
the
shP1-‐expressing
SupT1
cell
line,
which
targets
PSIP1.
In
contrast
to
a
previous
report,
no
change
in
the
ratio
of
unspliced
to
singly-‐
or
multiply-‐spliced
HIV-‐1
transcripts
were
detected
in
shH1-‐expressing
SupT1
cells,
suggesting
that
HTATSF1
does
not
function
as
a
splicing
cofactor
in
this
system.
A
slight
rebound
in
p24
levels
at
14
days
post-‐infection
was
accompanied
by
increased
HTATSF1
expression
and
a
decrease
in
the
percentage
of
cells
with
transgene
expression
in
the
population.
In
addition,
there
was
a
slight
decrease
in
shH1-‐derived
guide
strand
expression,
but
no
change
in
transcription
rates
of
the
htatsf1
gene,
suggesting
that
cells
within
the
population
with
shH1
expression
and
HTATSF1
suppression
may
have
a
growth
disadvantage.
Thus,
although
this
work
demonstrates
for
the
first
time
that
HTATSF1
functions
as
an
HDF
in
T
cell-‐derived
SupT1
cells,
it
may
not
constitute
a
viable
therapeutic
target.
A
second
objective
of
this
thesis
was
to
examine
the
feasibility
of
transcriptional
gene
silencing
(TGS)
of
HDFs
as
an
anti-‐HIV
strategy.
TGS
is
a
small
RNA-‐induced
gene
silencing
pathway
that
operates
through
chromatin
remodelling
with
the
potential
to
mediate
long-‐term
silencing
of
gene
expression.
Thus,
its
application
may
reduce
the
frequency
of
drug
administration
and
associated
toxicities.
Short
interfering
RNAs
(siRNAs)
targeting
the
htatsf1
promoter
were
able
to
reduce
target
mRNA
expression,
which
was
accompanied
by
decreased
htatsf1
transcription
rates
in
HEK293T
cells,
suggesting
silencing
via
a
TGS
mechanism.
The
htatsf1
silencing
inhibited
infectious
HIV-‐1
particle
production
from
TZM-‐bl
cells.
This
work
provides
proof
of
principle
that
TGS
induction
at
a
HDF
may
inhibit
HIV-‐1
replication.
siRNAs
targeting
the
ddx3x
promoter
did
not
induce
TGS.
To
examine
whether
gene
susceptibility
to
TGS
may
be
influenced
by
promoter
architectures,
49
promoter
features
were
examined
for
enrichment
in
genes
at
which
small
RNA-‐induced
TGS
has
been
reported.
Initially,
the
TGS
group
was
compared
to
a
random
set
of
2,000
promoters
and
then
all
other
promoters
in
the
genome.
To
control
for
gene
activation,
two
further
analyses
were
performed
comparing
the
TGS
group
features
to
those
from
promoters
active
in
the
THP-‐1
cell
line
and
housekeeping
genes.
Whilst
difficult
to
ascribe
differences
between
the
TGS
group
and
the
control
groups
to
anything
beyond
a
variation
in
the
proportion
of
active
genes
within
each
group,
there
was
enrichment
for
certain
promoter
features
that
are
independent
of
activity;
the
TGS
group
was
characterised
by
broad
transcription
start
regions,
high
CpG
content
and
a
single
expression
profile.
Moreover,
the
fraction
of
promoters
with
reported
non-‐coding
RNA
overlap
was
greater
in
the
TGS
group
than
the
control
groups.
Thus,
there
is
some
evidence
that
a
number
of
promoter
features
are
associated
with
TGS
susceptibility.
It
is
hoped
this
novel
analysis
will
facilitate
selection
of
future
TGS
targets,
including
HDFs.
In
summary,
the
work
presented
in
this
thesis
paves
the
way
for
development
of
improved
anti-‐HIV
therapies
involving
HDF-‐targeted
TGS-‐based
gene
therapies
that
mediate
sustained
inhibition
of
the
virus.
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169 |
Estudos das relações quantitativas entre a estrutura e atividade de uma série de inibidores da protease do vírus HIV-1 / Quantitative Structrure activity relationships for a series of HIV-1 protease inhibitors.Ferreira, Leonardo Luiz Gomes 01 March 2007 (has links)
A Protease do Vírus da Imunodeficiência Humana Tipo 1 (HIV-1 PR, EC 3.4.23.16) é um alvo macromolecular de grande importância no desenvolvimento de fármacos na terapia da Síndrome da Imunodeficiência Adquirida (AIDS). As maiores indústrias farmacêuticas do mundo concentram inúmeros esforços em estudos acerca desta proteína, que desde a introdução do saquinavir (Invirase®) na terapêutica em 1995, tem se mantido como um alvo fundamental para a descoberta de novos fármacos anti-HIV. A protease do vírus HIV-1 possui uma história rica de enorme sucesso no processo de descoberta e desenvolvimento de fármacos. A Química Medicinal moderna, de forte caráter multidisciplinar, fornece um arsenal de alternativas e estratégias racionais úteis no processo de planejamento de novos fármacos. Uma das tecnologias muito empregadas com sucesso é o estudo das relações quantitativas entre a estrutura e atividade (QSAR) para conjuntos de dados padrões. Os estudos de QSAR visam identificar e quantificar no complexo campo de modelagem as relações entre a estrutura e atividade de uma série de moléculas. Na presente dissertação de mestrado, foram realizados estudos de QSAR bi- (2D) e tridimensionais (3D) empregando, respectivamente, as técnicas holograma QSAR (HQSAR) e a análise comparativa dos campos moleculares (CoMFA), visando à geração de modelos preditivos para um conjunto de inibidores da protease do HIV-1. Os modelos gerados, associados às informações obtidas pelos mapas de contribuição 2D e de contorno 3D, são guias químico-medicinais úteis no planejamento de novos inibidores mais potentes e seletivos da protease do HIV-1. / The Human Immunodeficiency Virus Type 1 Protease (HIV-1 PR, EC 3.4.23.16) is a macromolecular target of great importance for the therapy of the Acquired Immunodeficiency Syndrome (AIDS). Major pharmaceutical companies around the world concentrate several efforts on studies concerning this enzyme, which since saquinavir (Invirase®) reached the market in 1995, has maintained its status as a fundamental target for anti-HIV drug discovery. HIV-1 protease has a rich history of enormous success in the drug discovery and development process. The strong multidisciplinary character of modern Medicinal Chemistry supplies an arsenal of useful rational strategies for the design of new drugs. One such technology is quantitative structure-activity relationships (QSAR), which has been successfully applied in a number of settings. QSAR studies aim to identify and quantify the relations between structure and activity of series of bioactive molecules organized within standard data sets. In the present master\'s dissertation, 2D and 3D QSAR studies were performed employing the hologram QSAR (HQSAR) and comparative molecular field analysis (CoMFA) techniques, respectively, in order to generate predictive models for a large set of HIV-1 PR inhibitors. The final models along with the information obtained from the 2D contribution and 3D contour maps should be useful in the design of new inhibitors with increased potency and selective within the chemical diversity of the data set.
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Representações sobre o período da primeira internação hospitalar na perspectiva de mulheres HIV positivas / Representations about the first hospitalization period in the perspective of HIV-positive womenGiraldi, Iara de Moura Engracia 05 May 2011 (has links)
Giraldi, I.M.E. Representações sobre o período da primeira internação hospitalar na perspectiva de mulheres HIV positivas, 2011 A aids foi relatada pela primeira vez em 1981 e atualmente a Organização Mundial da Saúde estima que aproximadamente 36 milhões de pessoas estejam infectadas pelo HIV em todo o mundo. No Brasil, entre as mulheres tem sido verificado o aumento da incidência de infecção a partir da segunda década da epidemia, indicando não apenas as dificuldades para oferecer respostas institucionais ao controle da epidemia, mas também, evidenciando as questões de gênero, em particular nas relações conjugais, como relações sexuais desprotegidas por falta de poder de negociação do preservativo e os comportamentos de risco adotados por seus parceiros, cuja assimetria provoca a vulnerabilização das mulheres à infecção. O adoecimento dessas mulheres leva a uma perspectiva preocupante, pois muitas vezes este adoecimento vem associado a responsabilidade dos cuidados de um parceiro e/ou de possíveis filhos infectados. Porém, quando os sintomas começam a aparecer, surgem ansiedade e medos que estavam aparentemente controlados. O processo de internação pode ocasionar reações que agravam o quadro dos pacientes internados. Neste sentido, este projeto teve por objetivos identificar, entre mulheres soropositivas para o HIV, algumas representações sociais sobre a primeira internação hospitalar motivada por manifestação de sintomas, adoecimento devido a fragilidade do sistema imunológico e/ou efeitos colaterais associados ao tratamento. Este estudo foi realizado com 10 mulheres soropositivas, com idade entre 32 e 46 anos, internadas numa unidade de tratamento específica - UETDI. A análise temática de conteúdo das transcrições de entrevistas individuais, semiestruturadas, audiogravadas foi sintetizada em Categorias e Subcategorias empíricas. Durante a internação, concretiza esta nova fase, sintomática, levando as participantes a encontrar novas formas de enfrentamento, representação do próprio corpo, novas perspectivas e, via contato com uma equipe adequada às suas necessidades e familiares, poder sair dessa hospitalização com novas possibilidades e representações de saúde. Por fim, pode-se indicar algumas reflexões acerca da complexidade da adesão do portador de HIV ao tratamento. / Giraldi, I.M.E. Representations about the first hospitalization period in the perspective of HIV-positive women, 2011 Aids was first reported in 1981; nowadays, the World Health Organization estimates that 36 million people are infected by the HIV worldwide. In Brazil, an increase of the infection\'s incidence has been observed in women since the epidemic\'s second decade. Such phenomenon indicates not only the difficulties for offering institutional responses in order to control the epidemics, but, also, it evidences genderrelated and more specifically conjugal questions, such as unsafe sexual relations occurring due to a lack of negotiation power for the use of condoms and risky behaviors adopted by partners, whose asymmetry leads to the increase of women\'s vulnerability to the infection. Women\'s process of sickening portrays a worrying panorama, for such process is associated to the responsibility for offering care to a partner or possible infected children. With the appearance of initial symptoms, though, anxiety and fears that were apparently under control arise, in contrast to an initial healthy state without weighty worries. In such context, the hospitalizing process can lead to reactions that aggravate the state of patients. The present study aimed to indentify some social representations of HIV-positive women regarding their first hospitalization due to symptom manifestation, immunologic fragility and/or treatment-related side effects. Ten women took part of the study, with ages between 32 and 46 years old, who were hospitalized in a specific treatment unity (UETDI). Thematic analysis of the recorded individual, semi-structured interviews\' contents was synthesized in empirical Categories and Subcategories. During hospitalization, a new, symptomatic stage becomes real, leading participants to find new strategies for coping, representing their own bodies, developing perspectives and, through the contact with health staff and family members, exiting the hospital with new possibilities and health-related representations. At last, some reflections are indicated regarding the complexity of adhesion to treatment process by people with HIV.
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