Graduating Senior Scholar Abstracts
Class of Summer 2018
Graduating Scholar Abstracts
- Effects of Vet TRIIP Yoga Program on Pain and Stress among Veteran Participants
Anjali Binoy, Caludia James, Bob Deschner, Dottie Goodsun, and M. Danet Lapiz-Bluhm, PhD, RN, MSCI
School of Nursing, University of Texas Health Science Center at San Antonio Vet TRIIP
Yoga is a mind and body practice with a 5,000-year history in ancient Indian philosophy (Feuerstein, 2012). The benefits of yoga for both physical and mental health have been supported by research; regular yoga practice yields benefits for pain and stress (e.g., Ross et al., 2010). Veterans face several risk factors for long-term mental health problems and reintegration to civilian life including posttraumatic stress disorder (PTSD), depression, chronic pain, mild traumatic brain injury (TBI), and other conditions which affect their quality of life and families (Lapiz-Bluhm & Peterson, 2014). A qualitative study exploring yoga in Veterans with PTSD symptoms indicated that they found benefits of yoga for mental stillness, body awareness, and social connection (Cushing et al., 2018). Interestingly, the Veterans identified barriers to the practice as being socially unacceptable, especially for men, and physically unchallenging (Cushing et al., 2018). With the purpose of honoring and empowering Veterans to create healthy, happy and productive civilian lives, Vet TRIIP (Veterans Team Recovery Integrative Immersion Process) Program provides a short-term multi-modality complementary integrative immersion program for veterans with PTSD and related symptoms (http://www.vettriip.org/). The services provided through Vet TRIIP are emotional freedom technique (EFT), aromatherapy, clothes-on therapeutic massage, qigong, Reiki, chiropractic care, meditation, reflexology, and acupuncture in a two-hour session, as well as yoga, the focus of this specific study. Veterans attended weekly yoga sessions held on Wednesdays at 11 am- 1pm, and at the end they complete a survey at the end of the yoga session asking them to rate their pain and stress at the start of session (Pain-In and Stress-In) and after the yoga session (Pain-Out and Stress-Out) using a Likert scale (1-10, with 1 as least pain and 10 as most pain). For this analysis, data from Veterans (N=92) who attended yoga sessions from January – June 2019 (Number of sessions = 305) were analyzed for differences in pain and stress scores before and after the yoga session. The mean Pain-In score was 4 (SD =1.9) while Pain-Out score was 2.3 (SD=1.6). The mean Stress-In score was 3.3 (SD=2.2) while mean Stress-Out score was 1.3 (SD=1.5). At some sessions, Veterans shared that the yoga improved their physical mobility (“I could move my leg more”) and increased their socialization (“If not for these sessions, I would not go out of the house”). The data show that the Vet TRIIP yoga program has been beneficial for Veterans; attendance of yoga sessions reduced their pain and stress levels. Yoga also improved their physical function and socialization. This program should be supported to allow more offerings to increase Veteran participation.
Triple Negative Breast Cancer Inhibition by Isothiocyanates from Cruciferous Vegetables
Simon Bruhn and Pothana Saikumar, PhD Department of Pathology;
UT Health San Antonio, TX
It is estimated that 41,760 women will die of breast cancer in 2019 in the United States, of which approximately 50% may be from Triple-negative breast cancer (TNBC), which lacks receptors for hormones estrogen and progesterone, and HER2 receptor. TNBC poses a unique challenge to current breast cancer treatments as it lacks these targets and has higher recurrence and metastatic rates. Only toxic chemotherapies are the current choice of treatment for TNBC, therefore, we explored new compounds with less toxicity to treat TNBC using in vitro cell culture models. We hypothesized that chemicals derived from cruciferous vegetables, such as cabbage, broccoli, brussels sprouts, cauliflower etc., will kill cancer cells by binding to the cysteine residues of physiologically important non-genomic protein targets. Organic compounds in crucifers have long been suspected to have anti-cancer activity. The drugs being tested in our study were Phenyl isothiocyanate (PITC), Benzyl isothiocyanate (BITC), and Sulforaphane (SFN), which are found in Cruciferous vegetables of the Brassicaceae family (commonly known as the cabbage family). In fact increased consumption of such vegetables have been shown to reduce the risk of developing breast cancer. The anticancer properties of these compounds were tested using cultures of highly aggressive triple negative breast cancer cells, MDA-MB-231. We measured the half maximal inhibitory concentration (IC50) of these compounds in inhibiting the cell viability of MDA-MB-231 cells using Cell Counting Kit 8 (CCK8) assay. Measurement of IC50 values of these three compounds after 24h of treatment, suggested that PITC (IC50: 7.5μM) is much more effective in reducing the cell viability of MDA-MB-231cells, followed by BITC (IC50: 26 μM) and then SFN (IC50: 54 μM). SFN is required about 7 x higher dose than PITC to achieve the same degree of inhibition of cell viability. Growth inhibition is further confirmed by measuring proliferation markers like pAkt and cell cycle inhibitors like p21 and p27 by Western blotting techniques. Our research is being continued to compare the effects of these drugs on normal cells (MCF 10 A) and TNBC cells (MDA-MB-231, HCC1937 and BT20) to assess the efficacy of isothiocyanates as anti-cancer drugs in the pre-clinical setting.
Maternal-Infant Stress Reactivity and Physiological Attunement in Prenatally Opioid-Exposed
Newborns
Victoria Davila, Kelly McGlothen-Bell, PhD, RN, IBCLC, Jacqueline McGrath, PhD, RN, FNAP,
FAAN, Elizabeth Brownell, PhD, Leticia Scott, MA, Natashia Bibriescas, MS, Danny Wang, MS, &
Lisa Cleveland, PhD, RN, CPNP, IBCLC, FAAN
The quality of early primary maternal-infant dyad relationships is critical to subsequent
childhood development. These relationships are supported by the reciprocal behavioral
responses within the dyad, known as physiological attunement. Evidence shows that prenatal
opioid exposure is an early life stressor for infants, possibly causing emotional, behavioral, and
cognitive dysregulation, as well as a disruption of maternal-infant physiologic attunement. While
researchers have examined physiological attunement in term and preterm infants, little is known
about the impact of prenatal opioid exposure on physiological attunement. Therefore, the
purpose of this study is to gain a better understanding of the impact of prenatal opioid exposure
on maternal-infant stress reactivity and recovery, and physiological attunement. Pregnant or
early postpartum women (N=220) and infants (N=220) shortly following birth are being enrolled
in this study. When infants are approximately six months old, mother-infant dyads are assessed
for stress reactivity, recovery, and physiological attunement using an established behavioral
experiment— the Still Face Paradigm (SFP). This experiment consists of three stages labeled play,
still face, and recovery. Each stage is approximately two minutes long, in which physiological
reactivity (heartrate variability, respiratory sinus arrhythmia, and salivary biomarkers) is noninvasively recorded at baseline, during each phase of the experiment, and following completion
of the SFP. Concurrently observational data is being recorded and coded for maternal-infant
interaction patterns. Following the experiment, mothers are asked to respond to a series of selfreport instruments where they indicate their past and present physiological states, including
childhood trauma, perceived stress, and anxiety. Presently, 43 mother-infant dyads have
completed all data collection. Further data collection and analyses are ongoing. The findings of
this investigation may provide a better understanding of the impact of prenatal opioid use on
early mother-infant interactions. The knowledge gained from this study may lead to the
development and implementation of interventions to support the early maternal-infant
relationship and improve stress response within this vulnerable population.
Involvement of RNA Binding Proteins Musashi1 and SERBP1 in Glioblastoma Development
and Treatment
Talia Delambre, Adam Kosti, Luiz O. F. Penalva, Ph.D
Greehey Children’s Cancer Research Center
Glioblastoma (GBM) treatment has not seen major improvements in decades with an average patient
survival of 15 months. Despite all genomic efforts led by the TCGA, recent therapeutic strategies based
on identified drivers and genomic alterations have been disappointing. To create new routes for therapy,
it is necessary to continue mapping new pathways contributing to gliomagenesis. In this regard, there is
growing evidence that RNA binding proteins (RBPs) are major contributors to expression alterations
affecting genes in signaling pathways and biological processes critical to GBM growth and response to
therapy. RBPs regulate gene expression via RNA processing, mRNA decay and transport and translation
and are differentially expressed in tumors at significantly higher levels than other gene families.
Importantly, distinct from previous therapeutic strategies that focused on a single proteins or pathways,
RBP targeting provides the opportunity of interfering with several cancer pathways at once.
We have established the stem cell regulator Musashi1 (Msi1) as a critical player in GBM and
medulloblastoma and as a marker of clinical outcome and response to therapy. To define Msi1 main
routes in tumorigenesis, we conducted a comprehensive genomic and functional analysis where we
mapped Msi1 target genes, evaluated its effect on expression, defined genes with high expression
correlation in GBM and evaluated its impact on tumor growth and cancer phenotypes. Our results showed
that DNA replication/repair and cell cycle/division are the processes affected by Msi1 the most. In our
model, Msi1 regulates directly and indirectly the expression of a network of genes, promoting cell cycle
progression and DNA replication. We propose Msi1 as a therapeutic target and we are working on
identifying inhibitors of this RBP.
We identified SERBP1 (Serpine1 mRNA binding protein 1) as a new oncogenic factor in GBM. SERBP1
is a member of the RG/RGG family of RNA binding proteins, which are known for their involvement in
neurological and neuromuscular diseases and cancer. As its name suggests, SERBP1 regulates the
expression of Serpine1 (PAI-1), a member of the serine protease inhibitor family that is often highly
expressed in tumors and implicated in tumor growth and metastasis. In respect to GBM, our study
indicates that SERBP1 uses two main routes to contribute to tumorigenesis. It promotes “stemness” by
repressing genes involved in neuronal differentiation and enhances the expression of genes driving
alternative metabolic pathways often used by cancer cells. Therefore, SERBP1 influences two critical
processes that are currently top priority in targeted therapy. Our project will not only deliver new
information on molecular mechanisms implicated in these processes but also an inhibitor to be potentially
used in the treatment of GBM and other malignancies affected by SERBP1.
SERBP1 is difficult to study because the beginning of the protein structure is disordered. In order
to investigate the structure of SERBP1, we require a shorter fragment which is structurally stable. Using
PCR based cloning, we produced a plasmid containing only the ordered part of the protein. We will
perform assays using this plasmid to compare the full and shortened lengths of SERBP1 to determine
whether the shortened fragment behaves differently than the full protein and whether they interact with
the same RNA binding motifs. The plasmid containing the shortened fragment of SERBP1 will also help us
discover the function of the disordered section of our protein of interest.
Ewings sarcoma: NMR investigations of the major fusion protein EWS-FLI1
Parth Ghawghawe, Xiaoping Xu, Ph.D., David Libich, Ph.D.
Department of Biochemistry and Structural Biology and Greehey Children’s Cancer Research Institute
Intrinsically disordered proteins are a class of proteins that do not have a fully defined
shape or ordered structure, but still carry out a function. Ewing sarcoma (EwS), caused by a
balanced chromosomal translocation commonly at the t(11;22)(q12;q24) location, is a malignant
bone tumor predominantly found in children and adolescents. EwS may be caused by multiple
different chromosome translocation combinations, each dependent on the location of the
translocation, but for this project, we focused on the major translocation EWS-FLI1. Due to the
unique nature of intrinsically disordered proteins, conventional structural methods such as X-ray
crystallography and cryo-electron microscopy can not be used to solve their structure.
Alternative methods, such as nuclear magnetic resonance (NMR), must be used to determine
their relative structure. The goal for this project was to create a sample of highly concentrated
protein to investigate by NMR spectroscopy.Due to the difficulty of expressing the full protein
formed from the fusion EWS-FLI1, the protein was further broken down into sections: Serine 194-
421 and Serine 218-445. These were chosen to include the functional portions of both the EWS
and FLI1 parent proteins, and because they are shorter protein lengths for improved expression.
Each section was inserted into vectors containing the solubility tags (maltose binding protein or
glutathione S-transferase), or a His6 tag, ultimately resulting in 6 constructs. Methods such as
polymerase chain reaction, gel electrophoresis, polyacrylamide gel electrophoresis, and DNA
sequencing, were all used to create and confirm the validity of each construct. Results from the
polyacrylamide gel electrophoresis showed that predominantly all of the expressed protein were
insoluble. Further testing demonstrated that when the target protein was expressed at 16 oC,
they were found to be more soluble than at 37 oC. To create the NMR sample, EWS-FLI1 218-445
was chosen because of its favorable expression in combination with the His6 tag/vector as the
target protein. A colony transformed with that vector/construct combination was grown in M9
medium and purified using a BiologicLP system with Immobilized Metal Affinity Chromatography
(IMAC) column purification method. Throughout the entire purification process, polyacrylamide
gel electrophoresis was used to test to see if the target protein was present. We observed that
the soluble fraction of the target protein, EWS-FlI1 218-445 with the histidine tag, would show
signs of degradation throughout the process, eventually leading to the majority of the protein
being lost to degradation. This was confirmed by the appearance of similar length bands
appearing during different stages of the purification, and as a result, unlikely lost due to human
error. For the insoluble fraction recovered from the purification, during high pH buffer exchange
to the low pH buffer, the protein would constantly precipitate, making it unstable in low pH
environments needed for NMR spectroscopy. We observed that the construct EWS-FLI1 218-445,
even with a solubility tag such as histidine, the soluble fraction of the target protein was still
extremely prone to degradation and a different approach must be used during purification. For
the insoluble fraction, a new method must also be developed to prepare the protein for the low
pH environment as the protein precipitated during buffer exchange, thus making it unusable for
NMR spectroscopy. Future efforts will focus on altering the expression and purification strategy
to improve recovery of soluble protein.
Effects of Gestational Diabetes and Metformin Exposure on Mouse Offspring Social Behaviors
Noah Hodge, Brandon Rice, Livia Ferreira, Yatzil Sanchez, John Yunger, Georgianna Gould
Rates of gestational diabetes mellitus (GDM) have increased in recent decades, and an
epidemiological association of GDM with an increased risk of autism was discovered. In light of
this, reports that the anti-diabetic drug metformin may control GDM better than the “goldstandard” insulin raises the exciting possibility that its use might also curb autism. Yet use of
metformin in pregnancy is still new, and it is readily transported across the placenta, so concerns
were raised about potential long-term adverse effects on offspring. In this study, mice with
pharmacologically induced GDM (by streptozotocin (STZ)), or high fat-high sucrose (HFHS) dietinduced GDM and their respective controls were treated with metformin (≈200 mg/kg/day) in
drinking water throughout pregnancy. Overall the STZ treated dams had fewer litters born than
dams from the other groups. Mouse offspring were tested for social interaction and social novelty
preference in three chamber choice tests when they were 7-8 weeks old. To increase scientific
rigor and reproducibility all behavior data was collected by students trained to measure a specific
behavior, and who were unaware of the treatment groups. In a prior published study, our lab
observed that males exposed during gestation to metformin engaged in less social interaction
than the vehicle control group. In this study we examined preference for social interaction and
social novelty through social sniffing of stranger mice, with time spent sniffing an empty cage or
known mouse at opposite end of the arena subtracted from the stranger sniffing time value. No
differences in male social interaction or preference were evident among treatment groups. For
males, the mean ± SEM preferences were: control 63.2 ± 7.5 sec, metformin 39±12.1 sec, STZ
53.3 ± 23.1 sec, and metformin 58.6 ± –––––14.5 sec (N = 4-14). Female offspring had similar
results. Likewise males and females exposed during gestation to HFHS diet ± metformin also
exhibited no differences between groups by measure of sniffing time. The reason for this
outcome was that small sample sizes in critical STZ and HFHS control groups, due to loss of
pregnancies and neonatal pups, did not provide critical data for detecting differences among
treatment groups. By contrast metformin appeared to increase the fertility of the mice in the
STZ and HFHS diet groups, and was reported in prior studies in fish to be an endocrine disrupter.
A follow up study is under way to obtain larger numbers for those groups with low sample sizes.
The offspring mice were also tested for repetitive behavior via a marble-burying test, and there
were no differences among treatment groups or sexes in this restrictive-repetitive behavioral
index, which averaged 5.6 ± 1.2 for male and 5.9 ± 1.8 for female offspring (N = 4 – 17).
Metformin-exposed male offspring were more dominant than controls (nonparametric p = 0.04,
N = 5), but offspring from GDM pregnancies did not exhibit this effect. While data collection is
still underway, these preliminary findings indicate that despite prior findings of adverse effects
of gestational metformin exposure on mouse offspring from non-GDM pregnancies on these
behaviors, here metformin exposure had no obvious adverse effects on offspring from GDM
pregnancies. Metformin enters the placenta via organic cation transporters and targets fetal
adenosine monophosphate-activated protein kinase (AMPK) in mitochondria, resulting in
metabolic responses which primarily help to control GDM. Under conditions of GDM, we predict
any potential adverse impacts of metformin on fetal development to be dissipated through
further research into the underlying mechanisms at work in this process.