Student projects

Internship project of Sheyla E. Cisneros Montalvo

Characterizing Sertoli cell markers during mouse testis development

Before coming to Turku I was a member of the Animal reproduction laboratory in my home university (Universidad Nacional Mayor de San Marcos, Lima, Perú) After arriving in Turku, first as an exchange student, I wanted to find a place where to learn more about reproductive biology and to develop my practical laboratory skills. Luckily I found such a place in the Male Reproductive Health, Developmental Disorders and Spermatogenesis group, leaded by MD, PhD, Professor Jorma Toppari in the department of Physiology, Faculty of Medicine, University of Turku.

During my internship I have been supervised by Juho Anti Mäkela and Emmi Rotgers, who have taught me a variety of techniques in lab work, such as cell culture, immunohistochemistry staining, genotyping, chromatin Immunoprecipitation (ChIP Assay), western blot, PCR and qRT-PCR.

The topic of my internship project was “Differential expression of Imagen1Sertoli cell markers in mouse testis development”. Sertoli cell are highly differentiated somatic cells in the testis that control testicular differentiation during development and support adult spermatogenesis. At present, most Sertoli cell markers are validated in humans, but since animal models, such as mouse, are extensively used also in the field of reproductive biology, it is important to validate these markers in different species. Furthermore, the definition and identification of the sequence of appearance of Sertoli cell markers during mouse testicular development may be important for understanding the role of their re-expression in testicular disorders with associated Sertoli cell dedifferentiation in adulthood. During the project we performed qRT-PCR and immunofluorescence staining of Wilms tumor 1 (WT1), anti-Müllerian hormone (AMH), keratine 18 (Krt18) and Vimentin (VIM) at different time points.

Picture: Immunofluorescence of VIM and WT1 of 10 weeks old of mouse testis.

 

Internship Project and Master’s thesis of Bettina Hutz

I used to work as a biological laboratory technician, most recently in a group that studied mitochondrial genetics and gene expression. I became interested in this field, so I was pleased to discover a group at the Department of Biochemistry at Åbo Akademi, which had just started implementing mitochondrial research into their profile, and where there is a demand for advanced imaging. After an informal meeting with the Group leader Thomas Nyholm and my supervisor Helen Cooper we soon discovered that I would fit into the team well, as I could focus on imaging and this way also significantly contribute to the group’s projects. Therefore we agreed on an internship project that I started immediately.

During my internship project I cultured different cell-lines, which my supervisor Helen Cooper had cloned and used for gene-expression experiments during her post-doctoral work at the Mitochondrial Biology Unit in Cambridge, UK. I have repeated some of her experiments as a kind of “validation” of the methods and conditions; to prove that they also work here in Finland, and in my hands. These experiments have included: Culturing and maintaining of wild-type and mutant transgenic cell lines, induction of gene expression, and immunofluorescence double-stainings of fixed cells at different time points after induction of gene expression. I performed all experiments with two differentially tagged genes, both wt and mutant, and also established more immunofluorescence staining protocols with some additional primary antibodies that allow more flexibility/combinations in parallel double-stainings. The cells were imaged with confocal scanning microscopy at the Cell Imaging Core (BTK Turku). 3D reconstructions of the images and colocalisation analyses were carried out using the BioImage XD software (Kankaanpää et al., 2012.) I was able to repeat the results that were obtained in Cambridge, and we got some more antibodies to work well. Therefore we are planning the next step, which will be my thesis project : The evaluation of  protein interactions in mitochondrial nucleoids before and after the knock-down of a mitochondrial inner membrane protein. The results will base exclusively on observations made with confocal microscopy and super-resolution microscopy at the Cell Imaging Core (BTK Turku).

Whenever I have some free-time between lectures, or on weekends and bank-holidays, I help my supervisor Helen with imaging some GMPVs (giant plasma membrane vesicles) from her other project, that examines lipid behaviour in integrin α2 clustering. It is a much more biophysical topic, but also involves confocal imaging.

Kankaanpää P, Paavolainen L, Tiitta S, Karjalainen M, Päivärinne J, Nieminen J, Marjomäki V, Heino J, White DJ. (2012) BioImageXD: an open, general-purpose and high-throughput image-processing platform. Nat Methods.

Picture: Co-localization study of a mitochondrial marker (red) and the transgene protein product fused to a reporter protein (GFP).

 

Master´s Thesis of Arsalan Khan

During my MSc. Biomedical Imaging, I have studied courses related to medical and preclinical imaging and I found them really interesting. So I have decided to work in MR imaging related project. I have applied for the research position through internet and got the thesis position in Cardiac Magnetic Resonance Imaging (CMRI) group of A.I. Virtanen Institute for Molecular Sciences at University Of Eastern Finland. In A.I Virtanen Institute for Molecular Sciences three animals MR scanners are installed, one is 7T broker and two are 9.4 T Varian magnets. Recently, PET scanner has been also installed in the institute. It’s a great place to work, if you want to work on pre-clinical MR imaging. The basic aim of my project was to measure and characterize the rotating frame relaxation time T1rho and also others (T2rho, TRAFF and T2) during myocardial infarction using magnetic resonance imaging (MRI). In the project, T1rho, T2rho, TRAFF and T2 have been measured in acute infarction. After imaging, the hearts were prepared for histology. Two different groups of mice have been tested. In first group invivo magnetic resonance imaging was done just after the infarction (1-2 hours) and then exvivo measurements have been done to verify results. After MRI, hearts were prepared for histology and I have used light microscope to take images. In second group invivo MRI has been performed just after infarction and after 7 days of infarction then exvivo MRI has been done. All the invivo and exvivo MRI measurements have been done using 9.4T Varian magnet.

Picture: T1r map of mouse heart after myocardial infarction

 

 Master´s thesis of Sergio Rondinelli

Since integrins trafficking is responsible for maturation of focal adhesions thus cell migration, cell development and mechanosensitivity in migratory cells. My project aimed to identify which mechanisms influence cells to migrate. We have identified that integrin α5β1 induce focal adhesions fast turnover and integrin αVβ3 stabilizes them. These are influenced by syndecan-4 adhesion receptor and by small GTPases activity as Arf-6 and syntenin. In fact cells that could not bind syndecan-4 at cell-matrix interface were not able to migrate and could not deliver integrin to cell-matrix interface. When cells were able to bind syndecan-4 receptor but Arf-6 and syntenin activity was silenced with siRNA, they could not migrate. I have used immunofluorescence assays and live-cell imaging to identify those processes.

Picture: 3i system of confocal spinning disk to perform live-cell imaging

 

Internship project of Elnaz Fazeli – Looking at the effect of bioactive lipid signaling molecules on intermediate filament structure

As an electrical engineer, I didn’t have any biology background. This internship helped me through understanding cell biology in practical matters. I learned how to do cell culture (cell preparation, cell treatment, staining and fixation), how to work with different fluorescence microscopes such as Confocal, STED and Brightflield Upright microscope and how to use my image processing theoretical skills with different processing software.

 

 

 

 

 

 

 

Master’s thesis of Mohamed El Missiry

My main interest is hematology, which is a branch of science highly dependent on biomedical imaging devices and applications. Currently I am doing my thesis on studying immunological responses of patients with chronic myeloid leukemia (CML) in comparison to healthy responses through flowcytometric analysis of T-lymphocytic subsets in respect to size, phenotype and function. T lymphocytes were grouped according to size and phenotype to small CD3+CD4+, large  CD3+CD4+, small CD3+CD4- and large CD3+CD4- cells.  Thereafter,  functional analysis was preformed through studying activated STATs (phosphorylated STATs) and correlating them to activated ERK and STATs of pathologic monocytic lineage of CML.  Different immunological responses could serve as  an indicator to leukemia versus healthy.

Picture: Flow cytometry data from T-lymphocytes.

 

 

 

 

eSci Research – Extracurricular activity of Muhammad Arsalan Khan and Mohamed El Missiry

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