Los Angeles Basin - California State University - MHIRT

Training Laboratories

Click the names of the Laboratory Directors below to read about the field of work and and find a link to their webpages.

United Kingdom

University of Cambridge

University of London

University of York

University of Oxford

Thailand

Chiang Mai University

Argentina

Instituto Fundación Leloir

University of Buenos Aires

National Institute of Infectious Diseases "Dr. C. G. Malbrán"

INBIRS (formerly National Reference Center for AIDS)

Insituto de Investigaciones Biotecnologicas "Dr. Rodolfo Ugalde"

Dr. Richard Farndale

Dr. Richard Farndale

University of Cambridge

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Collagen receptors and other collagen-binding proteins

This work relates to thrombotic disease, stroke and heart disease, conditions identified within the group of those where the most striking disparities are found.

My work centers on the investigation of interactions of the fibrous collagens with their receptors and other proteins. Focus was initially on the platelet collagen receptors, integrin alpha2beta1 and glycoprotein VI. This area has now been extended to include the indirect interaction between collagens and platelet glycoprotein Ib mediated by von Willebrand factor. The synthesis iof the Collagen Toolkits, libraries of peptides embracing the whole of the triple-helical domain of collagens II and III has enabled us to address the interaction with any collagen-binding species, including novel receptors, (DDRS and LAIRs) and other well-establsihed collagen-binding proteins such as fibronectin and SPARC.

We study the signaling pathways activated in human platelets by collagens of the blood vessel wall. These processes are fundamental to thrombosis and haemostasis, and our aim is to identify amino acid sequences in the collagens which recognize platelet receptors, to identify platelet receptors for collagen, and to understand the signaling consequences of their interaction. Again, this approach can be applied to other receptors in different cellular settings.

The project will form a part of the above program, and will be concerned with identifying signaling properties of collagen-derived synthetic peptides. This will allow us to locate platelet-activatory motifs with the collagen sequence. Measurements to be made may include the activation of phospholipase A2, of protein kinase C, and of tyrosine kinases. The work will involve SDS-PAGE, Western blotting and possibly immunoprecipitation. The laboratory is well-established: usually there will be 2 Ph.D. students and 6 post-doc researchers in the group, with other students working on different topics sharing facilities with us.

Dr. David Glover

Dr. David Glover

University of Cambridge

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Developmental Regulation of Cell Division in Model Organisms and in Cancer Cells

Minority groups suffer disproportionately from cancer, and disparities exist in both mortality and incidence rates.

Model organisms provide rapid insight into the mechanisms and control of cell cycle since they facilitate both genetic and cell free studies. Drosophila offers an excellent model for studying fundamental aspects of cell division cycle control and how these might differ between tissues throughout development. The organisation of the mitotic spindle and its function are also much closer to that of human cells than the yeasts, the other major genetic model for studying cell cycle control. The value of Drosophila as a model for studying cancer comes not only from our discoveries that many conserved cell cycle regulators are mis-regulated in cancer cells, but also that the cellular processes of the centrosome duplication cycle and of chromosome segregation go awry in tumour cells. We are able to use Drosophila as a developmental system in which to model these processes. Our genetical and cell biological studies on regulatory mitotic proteins in Drosophila have highlighted their multiple roles in mitotic and meiotic progression and how these can vary in different tissues. The group continues to identify components of these regulatory networks. We study their interactions both genetically and biochemically, and follow their behaviour in cells using both conventional and confocal microscopy on fixed and living specimens. As the mitotic and meiotic apparati are dynamic molecular machines, such time lapse imaging of the behaviour of individual molecular components is particularly valuable. The development of RNAi techniques enables us to study virtually any protein from the fly or human genomes. We have the capability to carry our whole genome RNAi screens in cultured Drosophila cells to identify regulatory proteins, and then study the roles of their human counterparts in normal and tumour cells.

Projects

Dissection of the multiple roles of Polo and Aurora kinases in mitotic progression using chemical genetics.

The evolutionarily-conserved mitotic kinases Polo and Auroras A and B have been shown to play multiple roles in mitotic progression. However, dissection of these functions has not been easily amenable using traditional genetics, due to the prolonged period before mutations produce a discernable phenotype. For this reason a chemical genetics approach has been proposed. This entails using small molecules that specifically target and perturb the function of the above kinases, acting immediately. The effects of inhibition of these kinases on mitotic progression will be examined in both normal and cancerous mammalian cell lines stably expressing fluorescently labelled components of the mitotic apparatus. Comparative studies will be applied to living Drosophila embryos, and cultured cells from larval brains and testes. In this way the effects of these compounds can be studied in concert with those of various mutations affecting cell cycle progression.

In vitro studies of microtubule nucleation by centrosomes

We have previously used an in vitro system to study the nucleation of microtubules by preparations of partially purified centrosomes. This has enabled us to study some of the roles of Polo kinase in recruiting and/or activating molecules associated with the minus ends of microtubules to increase the nucleating capacity of the centrosome on mitotic entry. We propose to further our understanding of the roles of protein kinases and protein chaperones using this in vitro system.

Protein kinases with mitotic functions

Mitotic progression is regulated through the concerted action of cycles of protein phosphorylation and dephosphorylation coupled to controlled proteolysis of key regulatory molecules. We discovered three of the principal regulatory mitotic protein kinases, Polo and the Auroras, through our work in Drosophila. We recently completed a screen of the entire fly kinome by knocking down the function of all of the protein kinases in the genome (239 enzymes) and examining the effects on cell cycle progression. This screen identified a number of protein kinases with mitotic roles that we now wish to study in detail using a combination of biochemical, genetic, and cell biological approaches.

 

Dr. Gary Martin

Dr. Richard Harvey

King's College University of London

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Dr. Harvey’s research is currently focused on studying molecular interactions between components of the bacterial cell envelope such as lipopolysaccharide and plasma membrane lipids, and membrane-active antimicrobial substances. To achieve this, his group uses a number of different experimental methods ranging from lipidomics and molecular spectroscopy to surface science techniques such as neutron reflectometry and diffraction. The overall aim of this work is to gain insight into physical mechanisms of drug tolerance in multi- resistant bacteria of clinical relevance, to aid the development of future anti-infectives.

 

Dr. Anne Duhme-Klair

Dr. Anne Duhme-Klair

University of York

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Siderophore-linked Drugs as Novel Antimicrobial Agents

The health disparities that afflict ethnic and racial minorities, as well as other underserved populations, are not limited to one or two disease categories. They apply to a broad spectrum of disease including infectious diseases. The problem presented by infectious diseases is compounded by the rise of antibiotic resistance strains that in some cases defy treatment with most if not all antibiotics. Infant mortality is augmented by infectious diseases, specially those caused by resistant strains and mortality is also increased in patients that are more susceptible to them as consequence of the presence of another disease such as AIDS, cancer or diabetes.

The problem of bacterial resistance to antibiotics is now so severe that it has been described as a crisis by the World Health Organisation. It is therefore important to find novel ways of combating the development of resistance. As iron is a key growth factor for most pathogenic bacteria, their survival depends on the ability to acquire Fe(III) from the host. To remove the Fe(III) from the mammalian Fe-binding proteins, bacteria produce and secrete high-affinity iron chelators, so called siderophores. These siderophores scavenge the iron by forming extremely stable Fe(III)-complexes.

Our approach is to use a 'Trojan Horse' strategy. This approach involves attaching the antimicrobial agent to a siderophore with the desired effect being that the antibiotic is taken up along with the siderophore when the bacterium is sequestering iron. If the bacteria mutate to block the entry of the siderophore-linked antibiotic, they will suffer from iron starvation and hence die. In addition, it has recently been recognised that multidrug efflux pumps select and bind hydrophobic drugs within the lipid bilayer of the membrane, the utilisation of the Fe-uptake system to transport drugs into the cell could bypass efflux pumps. The aim of this project is to synthesise a selection of siderophore-linked drugs, to study their iron-binding properties and to test their antibacterial activity against a panel of pathogenic organisms.

Techniques involved: The work will involve synthetic organic chemistry, UV/vis-spectroscopy and basic microbiology.

Background of the student: Some experience in synthetic chemistry and knowledge of basic microbiological techniques would be of advantage.

Dr. Anthony Wilkinson

Dr. Anthony Wilkinson

University of York

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Structural studies of gankyrins

Minority groups suffer disproportionately from cancer, and disparities exist in both mortality and incidence rates. This is a project designed to characterize Gankyrin, a protein overexpressed in certain types of cancer cells.

Gankyrin is a protein which is overexpressed in certain types of cancer cells. It induces anchorage independent cell growth and tumour formation in certain mammalian cell cultures. The protein consists of six ankyrin repeat motifs. The ankyrin repeat is one of most common protein motifs. It consists of an ~33 residue sequence which in different proteins is repeated as few as two, and as many as 20 or more, times. The ankyrin repeat domains appear to be important in mediating protein-protein interactions. Our aim in the current project is to determine the structure of gankyrin by X-ray crystallography. We have E. coli clones which direct high level expression of recombinant protein. The first step will be to purify the protein by column chromatography methods and determine its subunit structure by analytical ultracentrifugation. The main aim is to grow crystals of gankyrin suitable for X-ray analysis. Should crystals appear we will attempt to solve their structure using diffraction methods.

Methods used: Growth of bacterial cultures, protein purification by column chromatography, gel-electrophoresis, analytical centrifugation, crystallisation.

Background of Student: The student should have experience in basic chemistry and biochemistry. Some knowledge of molecular biology would be an advantage.

 

Dr. Kieran Clarke

Dr. Kieran Clarke

University of Oxford

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Cardiac stem cells to prevent heart failure – An MR study

Heart disease has been identified as relevant within the realm of health disparities. The "Strategic Research Plan and Budget to Reduce and Ultimately Eliminate Health Disparities - Volume I," published by the National Institutes of Health indicates that coronary heart disease mortality is 20 percent higher for African Americans than Whites and the incidence of stroke is disproportionately high in African Americans. See (http://ncmhd.nih.gov/our_programs/strategic/pubs/VolumeI_031003EDrev.pdf)

A promising, novel approach to the treatment of myocardial infarction and the prevention of heart failure is cell grafting in the damaged myocardium. The aim of this project is to determine whether cardiac muscle and vasculature can be formed from cardiac-derived stem cells (CSCs) to restore function and metabolism after coronary occlusion. CSCs will be isolated, expanded, characterized and transplanted into the chronically infarcted rat heart in vivo. The effects of the transplanted CSCs on heart function will be monitored non-invasively using magnetic resonance imaging (MRI). We propose to determine the optimal method of cell delivery and timing of cell implantation post infarction. We will also use magnetic resonance spectroscopy (MRS) to determine the extent to which the implanted stem cells prevent changes in myocardial energetics, mitochondrial function and substrate metabolism that are associated with remodelling in the failing heart. This project will combine stem cell experience with cardiac MR, physiological and biochemical expertise.

Our hypothesis is that functional cardiomyocytes, endothelial cells and vascular smooth muscle cells can be derived from primitive cells found in the heart and that these cells have the potential for treating the infarcted and/or failing heart. The student will isolate, expand and characterize the CSCs. They will then determine the therapeutic potential of these autologous CSCs in vivo in a rat chronic infarct model of heart failure by monitoring their effectiveness in controlling infarct expansion and remodelling, using MRI and MRS, and to determine the optimal method and timing of cell delivery.

Figure 1

The student would prepare cardiac stem cells, grow them on biomembranes, transduce them with GFP, label them with iron oxide particles and introduce the cells into the area of the infarct. Control groups will be injected with culture medium or non-functional control cells. Infarcts will be produced using left anterior descending coronary artery ligation and stem cells will be either administered via tail vein infusion or injected into the border zone of the scar tissue, with or without a scaffold. The infarct size and global and regional cardiac function and morphology during remodelling and failure will be determined, regularly and non-invasively over the following weeks, using in vivo MRI. After several weeks, rat hearts will be isolated for histology and immunohistochemistry or perfusion in the MR spectrometer for measurement of energetics and pH using 31P MRS. Cardiac output and efficiency will be determined in the isolated working rat heart. Transplanted cells will be tracked using iron oxide with MRI and by their expression of GFP using immunohistochemistry to identify the expression of marker plus cardiac-specific proteins in the transplanted cells.

In this project, the student would be working with postdoctoral fellows who routinely use the methods involved. Consequently, they would quickly learn the experimental techniques and should complete part of the project within the time allowed.

 

Dr. David Sherratt

Dr. David Sherratt

University of Oxford

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Mechanisms and functions of recombination in relation to dissemination, establishment, and persistence of antibiotic resistance genes.

Many of the diseases relevant to "health disparities" include as a risk factor the possibility of secondary bacterial infections. The current epidemic of superbugs (bacteria resistant to multiple antibiotics) increases the cost of treatment and the mortality rates. Furthermore, another issue relevant to "health disparities" as it infant mortality is greatly affected by the rise in the number of multiresistant pathogenic bacteria.

Our research search concerns the molecular mechanisms of various types of recombination processes, and how these mechanisms relate to biological functions. In this project, those functions are related to bacterial resistance to antibiotics. It is known that an important strategy bacteria utilize to resist antibiotics is through acquiring "resistance genes" usually by plasmid transfer. Plasmids posses a number of mechanisms to ensure their stable maintenance within the host bacterial cells. One of these mechanisms consists of resolution of plasmid dimers, which are formed through recombination events in the cell. Dimerization of plasmids leads to multimer formation, a known cause of plasmid instability. Xer is a site-specific recombination system that catalyzes conversion of plasmid dimers into monomers, ensuring plasmid stability.

The broad objective of this project is the understanding of the molecular mechanisms and interactions of the Xer recombination proteins and the plasmid under the various conditions the host bacterial pathogen may encounter through its life cycle. Upon entering a host, the pathogens' surroundings change dramatically, and they must have systems that enable them to grow or survive. Plasmids harbored by these bacterial cells carry genetic elements to be stably maintained in the various environments, and the molecular mechanisms may have to adapt to these changes. For example, a number of bacterial species cause a substantial amount of hospital-acquired urinary tract infections, pneumonia, septicemias, meningitis, and soft tissue infections. The niches occupied in these diverse infections may present different challenges to plasmids for stable inheritance

Dr. Kriengkrai Srithanaviboonchai

Dr. Kriengkrai Srithanaviboonchai

Chiang Mai University

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Project 1: People living with HIV

Students training in this location will carry on Public Health projects related to social attitudes, treatments, access to treatment, and compliance of people infected with HIV. Issues to be studied are: a) the community stigma and social support and their association with HIV treatment adherence among infected people of different social strata, b) identification of stigma and socio-economic status and their effects on patients’ satisfaction at health care facilities, c) susceptibility of HIV infected patients to diseases other than those due to impairment of the immune system such as cardiovascular problems and hypertension.

Project 2: Incidence of cancer in rural and factory workers

Another subject of work at the Department of Community Medicine, Chiang Mai University will be the high incidence of cancer in rural populations due to poor air quality as well as in factory workers due to the conditions of labor. Students will explore and propose solution to issues such as the conditions of labor in local factories, the utilization of pesticides by farmers, as well as cultural habits such as the “slash and burn” in the fields that has been done for centuries and results in high levels of lung cancer incidence.

 

Dr. Angeles Zorreguieta

Dr. Angeles Zorreguieta

Instituto Fundación Leloir

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Antibiotic resistance Drug resistant bacteria are commonly isolated from healthy persons as well as from patients with community-acquired infections in developing countries. Furthermore isolation of multiresistant strains from intrahospitalary infections have become the norm rather than the exception. This is due to a number of factors that include but are not limited to the high incidence of infectious disease, excessive clinical use of antibiotics (a problem also in industrialized countries), and their misuse by the public (in many developing countries antibiotics can be purchased without prescription). The problem is serious because as antibiotic resistance is making existing treatments less effective, development of new antibiotics is slowing down. We are working to characterize mechanisms of dissemination of genes coding for antibiotic resistance and to develop strategies to extend the life of existing antibiotics.

A number of diseases that disproportionately affect some populations are known to weaken the immune system increasing the susceptibility to bacterial infections. The rise of antibiotic resistance complicates their treatment.

Dissemination of genes coding for antibiotic resistance: we have recently characterized an integron that harbors clinically relevant resistance genes such as blaCTX-M -2, which codes for CTX-M-2, an extended-spectrum beta-lactamase from the growing family enzymes that preferentially hydrolyze cefotaxime over ceftazidime; and aac(6')-Ib, a gene encoding resistance to several important aminoglycoside antibiotics. This integron has been found associated to transposition genes making it potentially mobile.

Extending the life of antibiotics: development of inhibitors of enzymes that degrade antibiotics has been very useful to extend the life of these drugs. For example, clavulanic acid, a beta-lactamase inhibitor produced from Streptomyces clavuligerus, extends the activity of certain beta-lactam antibiotics against bacteria which owe their resistance to the production of beta-lactamases. We are developing strategies to interfere with expression of enzymes that confer resistance to non-beta-lactam antibiotics.

Dr. Daniela Centrón

Dr. Daniela Centrón

Department of Microbiology, School of Medicine, University of Buenos Aires

Dissemination of resistance genes in pathogenic Enterobacteriaceae. Impact on infant mortality.

One of the most efficient and versatile molecular mechanisms behind the rapid dissemination of antibiotic resistance genes is through integrons, genetic elements that act like natural expression vectors capable to pick up new resistant genes and express them at high levels. The students will be trained in the molecular biology and epidemiology of multidrug resistant enterobacterial clinical isolates, including but not limited to K. pneumoniae, that cause high mortality in hospitalized patients and neonatal wards. The training will include the characterization of integrons, their localization in the chromosome or plasmids, and their distribution among strains from different hospitals in Argentina.

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Dr. Alejandro Petroni

Dr. Alejandro Petroni

National Institute of Infectious Diseases

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Epidemiology of bacterial infections: resistance to antibiotics. Quinolone surveillance and mechanistic studies.

Quinolones are broad-spectrum synthetic antibiotics highly efficient for the treatment of enterobacterial infections. Unfortunately, they are not exempt from the current multidrug resistance crisis. Unlike with other antibiotics, the complexity and multiplicity of the mechanisms that decrease quinolone susceptibility make their clinical diagnosis very difficult because of the complementation, overlapping and masking effects. Furthermore, the recently discovered plasmid-mediated quinolone resistant mechanisms produce a slight increase in resistance leading the clinical laboratories to sometimes falsely categorize the bacteria carrying these plasmids as “susceptible”, with the consequent risk of therapeutic failure and the misidentification by the antimicrobial resistance surveillance systems. In this project the student(s) will participate in a national study involving public hospitals, which serve the less affluent population, to properly characterize the mechanisms that decrease quinolone susceptibility in clinical enterobacteria and to estimate their prevalence. The clinical objective of this project is to improve the diagnosis of these mechanisms at the level of the clinical laboratory and to implement a continuous surveillance system to follow their evolution.

Dr. Manuel Gomez Carrillo

Dr. Manuel Gomez Carrillo

National Reference Center for AIDS

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Epidemiology of HIV in diverse populations

This center studies HIV/AIDS from numerous points of view. Students with different backgrounds will be able to take advantage of training in this location. There is Public Health aspect that studies problems such as the assessment of the likelihood with which populations from big metropolitan cities or small cities will take preventive action as a function of the information provided to them through the Health Belief Model, the evaluation of the perceptions among sex workers from different regions about the preventive value of condom use, or the assessment of barriers to effective treatment of HIV patients that contract the hepatitis C virus, which seems to accelerate of the course of HIV disease but at the same time in these patients follows a faster course from hepatitis to cirrhosis and death. There is also a basic research division in the center that offers possibilities to our students to work on genomics and molecular epidemiology of HIV in populations at risk, such as the determination of genotypic and phenotypic characteristics of groups or populations that show less response to antiretroviral treatment.

Dr. Diego Comerci

Dr. Diego Comerci

Instituto de Investigaciones Biotecnologicas "Rodolfo Ugalde"

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Nanosensors and bionanoconsumables platform for Point of care –-POC- detection of infectious diseases (NANOPOC)

This project aims to consolidate and extend the capabilities of a nano-micro-biotechnology platform to generate nanosensors and bionanoconsumables for in situ detection of infectious diseases that affect human and animal health. This platform will include human resources, knowledge, infrastructure, regulation, portfolio of services and necessary public-enterprise base in the short and medium term, generating commercial products combining nano - bio and microtechnologies.

Through the use of nanoparticles and films of self-assembled monolayers (SAMs) bearing specific recognition of bio, nanosensors will be developed for high sensitivity and specificity. Moreover, the nano-scale reduction of the parent and / or supports will not only significantly increase the sensitivity but will also contribute to lower detection limits and times of the test. These bionanosistems will be combined with microelectronic devices such as lab-on-a-chip and optomicroelectronics like MEMS or MOEMS, which will produce highly innovative detection systems, both regionally and globally. The combination of technologies will allow the development of multiparameter diagnostic systems POC (point of care).

In this market there are no national or regional portable diagnostic systems, low cost, easy to use and able to detect a range of infectious diseases. Therefore the proposed technology platform will create a market niche that expresses itself a social necessity. During the first phase, we will focus on developing recombinant antigens such as proteins, glycoconjugated and monoclonal antibodies for diagnosing E. coli STEC (HUS), Chagas Disease and Brucellosis.

This platform shall be such that the private sector, the government in its various bodies and the scientific-technical system in general will have access to infrastructure, multidisciplinary expertise for the application of nanotechnology in general and the nano-bio-synergy microtechnology in particular for the development of diagnostic systems and advanced detection for a wide range of diseases and / or substances of commercial and social interest.

CSUF - California State University Fullerton