Dissecting the Role of Kinase Isoforms Using New Tools for Activation and Inactivation of Kinases in vivo

The Src family kinases (SFKs) are comprised of nine highly homologous members that participate in various signaling pathways, to regulate events such as proliferation, differentiation, cell migration, adhesion, and survival. While Src has been intensively studied, less is known about other family members. Unfortunately, similarities in the structure and regulation of SFKs have made it difficult to dissect their unique functions. Differential expression of individual SFKs have been found in various cancer cell lines, yet their functional specificity remains largely unknown. Previous strategies to investigate SFK specificity, such as overexpression and use of non-selective inhibitors, have been hampered by genetic compensation or lack of specificity. Therefore, it was imperative that new tools be developed in order to identify specific roles for individual SFKs. This dissertation describes two strategies to manipulate the activity of SFKs and to reveal novel information about their functional specificity in cell motility. The first study I performed used a small molecule to specifically activate each SFK isoform and to elucidate possible mechanisms contributing to the distinct activation phenotypes of Src and a Src family member, Fyn. In this study, new computational methods were developed to automatically analyze the dynamics of morphological changes. In the second study, I used light to manipulate Src activity. These strategies not only provide absolute control of specific Src family members, but also allow for spatial control of kinase activity. Additionally, these strategies utilize the same conserved site in the catalytic domain of the kinase and can be broadly applied to other tyrosine kinases.Doctor of Philosoph

Causal Mediation Analysis for Difference-in-Difference Design and Panel Data

Advantages of panel data, i.e., difference in difference (DID) design data, are a large sample size and easy availability. Therefore, panel data are widely used in epidemiology and in all social science fields. The literatures on causal inferences of panel data setting or DID design are growing, but no theory or mediation analysis method has been proposed for such settings. In this study, we propose a methodology for conducting causal mediation analysis in DID design and panel data setting. We provide formal counterfactual definitions for controlled direct effect and natural direct and indirect effect in panel data setting and DID design, including the identification and required assumptions. We also demonstrate that, under the assumptions of linearity and additivity, controlled direct effects can be estimated by contrasting marginal and conditional DID estimators whereas natural indirect effects can be estimated by calculating the product of the exposure-mediator DID estimator and the mediator-outcome DID estimator. A panel regression-based approach is also proposed. The proposed method is then used to investigate mechanisms of the effects of the Covid 19 pandemic on the mental health status of the population. The results revealed that mobility restrictions mediated approximately 45 % of the causal effect of Covid 19 on mental health status

Hydrothermal–galvanic couple synthesis of directionally oriented BaTiO3 thin films on TiN-coated substrates

AbstractBaTiO3 films were synthesized on TiN-coated Si substrate below 100°C by a hydrothermal–galvanic couple technique in barium contained alkaline solutions. X-ray diffraction and electron backscatter diffraction results show that the BaTiO3 thin films were directionally oriented grown on the TiN/Si substrates, i.e., (111) BaTiO3 over (111) TiN. The surface morphologies revealed that BaTiO3 nucleated and grew over the TiN surface with a single layer. From kinetic analyses, the growth rates of BaTiO3 films prepared by the hydrothermal–galvanic couple technique were faster than a hydrothermal method. The galvanic effects were confirmed by investigating the induced currents and energies. The galvanic currents were generated and controlled by both the dissolution of TiN and the formation of BaTiO3. The output electric energies increased rapidly with the reaction time and leveled off at the full coverage of BaTiO3

Surface exciton polaritons in individual Au nanoparticles in the far-ultraviolet spectral regime

All surface-excitation studies of Au in the past focused on the well-known 2.4 eV surface plasmon polariton in the visible spectral regime. The existence of surface exciton polaritons is believed to be pristine to the spectral regimes, showing strong excitonic absorptions. The presence of surface exciton polaritons in far-UV in Au (≥10 eV), where the optical and electronic properties of Au are dominated by broad interband transitions that display characters of rather weak and diffused excitonic oscillator strengths, is not expected and has never been discussed. Re-examining the reports of Yang and using electron energy-loss spectroscopy with a 2Å electron probe in aloof (optical near-field) setup and real-space energy-filtered imaging, we firmly establish the existence of surface exciton polaritons in individual Au nanoparticles in the far-UV spectral regime. These results indicate that surface exciton polaritons indeed can be excited in weak excitonic onsets in addition to their general believing for the sharp excitonic oscillations. Our experimental observations are further confirmed by the theoretical calculations of electron energy-loss spectra. The unmatched spatial resolution (2Å) of the electron spectroscopy technique enables an investigation of individual nanomaterials and their surface excitations in aloof setup. The surface exciton polaritons in individual Au nanoparticles thus represent an example of surface excitations of this type beyond the visible spectral regime and could stimulate further interests in surface exciton polaritons in various materials and applications in novel plasmonics and nanophotonics at high energies via manipulations of the associated surface near fields. © 2008 The American Physical Society.This work was supported by the National Science Council of Taiwan under Projects No. NSC94-2120-M-002-016 and No. NSC94-2119-M-002-025.Peer Reviewe

Overexpression of α-enolase correlates with poor survival in canine mammary carcinoma

<p>Abstract</p> <p>Background</p> <p>α-Enolase (ENO1) is a key glycolytic enzyme implicated in the development of many human cancers including breast cancer. Increased expression of ENO1 has recently been reported in estrogen (ER)-positive human breast cancer patients. The present study examined the expression of ENO1 and assessed its significance in canine mammary carcinoma.</p> <p>Results</p> <p>Immunohistochemical staining was employed to investigate the expression of ENO1 in 82 cases of canine mammary tumor (32 benign tumors and 50 carcinomas). Quantification of immunohistochemistry was carried out using Quick score and the results showed cytoplasmic ENO1 overexpression in 9 of the 50 carcinomas (18%). Overexpression of ENO1 correlated significantly with shorter cause-specific survival (P = 0.019), but was not associated with ER positivity in canine mammary carcinoma.</p> <p>Conclusions</p> <p>Our findings suggest that overexpression of ENO1 may be used as a prognostic marker for poor outcome in canine mammary carcinoma.</p

An optogenetic tool for the activation of endogenous diaphanous-related formins induces thickening of stress fibers without an increase in contractility: Photo-activation of Diaphanous-related Formins

We have developed an optogenetic technique for the activation of diaphanous related formins. Our approach is based on fusion of the Light-Oxygen-Voltage 2 domain of Avena sativa Phototrophin1 to an isolated Diaphanous Autoregulatory Domain from mDia1. This “caged” diaphanous autoregulatory domain was inactive in the dark, but in the presence of blue light rapidly activated endogenous diaphanous related formins. Using an F-actin reporter we observed filopodia and lamellipodia formation as well as a steady increase in F-actin along existing stress fibers, starting within minutes of photo-activation. Interestingly, we did not observe the formation of new stress fibers. Remarkably, a 1.9 fold increase in F-actin was not paralleled by an increase in myosin II along stress fibers and the amount of tension generated by the fibers, as judged by focal adhesion size, appeared unchanged. Our results suggest a decoupling between F-actin accumulation and contractility in stress fibers and demonstrate the utility of photoactivatable diaphanous autoregulatory domain for the study of diaphanous related formin function in cells

Elevated Krüppel-like factor 4 transcription factor in canine mammary carcinoma

<p>Abstract</p> <p>Background</p> <p>Krüppel-like factors (KLFs) are critical regulators of biological and physiological systems and have been extensively studied for their roles in cell proliferation, differentiation and survival in the context of cancer. Among the KLFs, KLF4 is highly expressed in human breast cancers and plays an oncogenic role. The present study examined the expression of KLF4 and assessed its significance in canine mammary carcinoma.</p> <p>Results</p> <p>Immunohistochemistry was employed to investigate the expression of KLF4 in 142 cases of canine mammary tumor. 75 of the 142 (52.8%) cases were histologically confirmed as mammary carcinoma. Quantification of immunohistochemistry was carried out using Quick score which multiply the staining intensity by the percentage of positive cells. High KLF4 expression was identified in 44 of the 75 (59%) dogs with mammary carcinoma and none in the benign cases. High KLF4 expression occurred only in the tumor cells and not the adjacent normal cells in mammary carcinoma (P < 0.001). Moreover, the high expression level of KLF4 expression was statistically associated with poor grade, late stage, histological subtypes of simple and complex carcinoma, and shorter 24-month survival. The Kaplan-Meier survival analysis also indicated that dogs with high nuclear KLF4 expression had a significantly shorter survival than those with low/moderate KLF4 expression (P = 0.011).</p> <p>Conclusions</p> <p>KLF4 is highly and frequently expressed in canine mammary carcinoma and correlates with a more aggressive phenotype.</p

Sequence variants of the aging gene CISD2 and the risk for Alzheimer's disease

Background/PurposeThe CISD2 gene has been related to life span control and mitochondrial dysfunction in animals. In addition, inhibition of mitochondrial enzymes due to an accumulation of beta-amyloid peptide has been related to Alzheimer's disease (AD). This study aimed to explore the association between sequence variants of the CISD2 gene and risk for AD, which has not been explored previously.MethodsThis was a case–control study involving a total of 276 patients with AD who were recruited from three teaching hospitals in Taiwan from 2007 to 2010; 460 controls were recruited from elderly individuals attending for health check-ups and volunteers in the hospital during the same period of time. All participants were aged 60 years or older. Two haplotype-tagging single nucleotide polymorphisms (htSNPs), rs223330 and rs223331, were selected from the CISD2 gene to test the association between their polymorphisms and the risk for dementia, and how ApoE ɛ4 status, sex, hypertension, and type 2 diabetes mellitus might modify this association.Resultsrs223330 variant carriage was not associated with risk for AD [TT versus CC: adjusted odds ratio (AOR) = 0.98, 95% confidence interval (CI) = 0.59–1.62; TC versus CC: AOR = 0.72, 95% CI = 0.47–1.11]. Similar findings were observed for rs223331 (AA versus TT: AOR = 1.12; AT versus TT: AOR = 0.99). In addition, hypertension significantly modified the association between rs223331 and risk for AD (p = 0.005).Three common haplotypes (with a frequency of 99.8%) were observed for CISD2. Common CISD2 haplotypes were not associated with the risk for AD.ConclusionOur findings suggested that CISD2 htSNPs are not associated with AD risk

Serotonin receptor HTR6-mediated mTORC1 signaling regulates dietary restriction-induced memory enhancement

Dietary restriction (DR; sometimes called calorie restriction) has profound beneficial effects on physiological, psychological, and behavioral outcomes in animals and in humans. We have explored the molecular mechanism of DR-induced memory enhancement and demonstrate that dietary tryptophan-a precursor amino acid for serotonin biosynthesis in the brain-and serotonin receptor 5-hydroxytryptamine receptor 6 (HTR6) are crucial in mediating this process. We show that HTR6 inactivation diminishes DR-induced neurological alterations, including reduced dendritic complexity, increased spine density, and enhanced long-term potentiation (LTP) in hippocampal neurons. Moreover, we find that HTR6-mediated mechanistic target of rapamycin complex 1 (mTORC1) signaling is involved in DR-induced memory improvement. Our results suggest that the HTR6-mediated mTORC1 pathway may function as a nutrient sensor in hippocampal neurons to couple memory performance to dietary intake

Women with endometriosis have higher comorbidities: Analysis of domestic data in Taiwan

AbstractEndometriosis, defined by the presence of viable extrauterine endometrial glands and stroma, can grow or bleed cyclically, and possesses characteristics including a destructive, invasive, and metastatic nature. Since endometriosis may result in pelvic inflammation, adhesion, chronic pain, and infertility, and can progress to biologically malignant tumors, it is a long-term major health issue in women of reproductive age. In this review, we analyze the Taiwan domestic research addressing associations between endometriosis and other diseases. Concerning malignant tumors, we identified four studies on the links between endometriosis and ovarian cancer, one on breast cancer, two on endometrial cancer, one on colorectal cancer, and one on other malignancies, as well as one on associations between endometriosis and irritable bowel syndrome, one on links with migraine headache, three on links with pelvic inflammatory diseases, four on links with infertility, four on links with obesity, four on links with chronic liver disease, four on links with rheumatoid arthritis, four on links with chronic renal disease, five on links with diabetes mellitus, and five on links with cardiovascular diseases (hypertension, hyperlipidemia, etc.). The data available to date support that women with endometriosis might be at risk of some chronic illnesses and certain malignancies, although we consider the evidence for some comorbidities to be of low quality, for example, the association between colon cancer and adenomyosis/endometriosis. We still believe that the risk of comorbidity might be higher in women with endometriosis than that we supposed before. More research is needed to determine whether women with endometriosis are really at risk of these comorbidities