PhD Portfolio - 2022

As you walk in the hallways of BMC, it is amazing how many internationals you see from all cultures and places. It is inspiring to see everyone working so well together.

Unfortunately, few struggles and obstacles come with being international: The migration issues!!!

I have been lucky to be able to join the lab as a visiting student before transition and becoming a PhD student. While I was on a scholarship, I was on a visiting residence permit, but I had to change that to a student residence permit when my PhD studies started.

The process has been quite challenging as I was asked to leave the country for this switch. Now, I was lucky to still have a permit to go to Germany, but if I did not, I cannot begin to think what I could do. I am from Gaza, Palestine and going back there is not possible at the time.

Luckily, things worked out and I started my position. However, I think the HR department at the University should reflect on how much preparation they need when they hire an international verses hiring a local. There could have been a lot of things improved in that regard.

After I had completed my bachelor’s degree in biochemistry, I was not sure about my career goals and what direction I should take. Thus, I started examining the different options. I ended up joining a research group where I learned a lot of techniques and my research interest has started to grow. Then, I knew that I wanted a career in academic research. Thus, doing a PhD is a step in the right direction for a career as an academic researcher. I chose to follow the PhD program at Lund University because of my research interests. I have been in the field of lung fibrosis for the past two years and I am happy that now I can pursue a PhD in it now.

Honestly, there is not much alternatives I would see myself in at this point. I enjoy doing academic research and I feel very confident with the choices I have made. But to reflect on the “What if?” question. There are few alternatives I could have been doing now. My original educational track was to pursue medical school and eventually train as a physician. To this end, even after I finish the PhD program, I may still pursue this goal. Alternatively, I could have been working in a pharmaceutical company as a technician or in project management. If I had chosen this path, I may have been much happier with the time spent on the job and the quality of my life outside the professional brackets. However, I am sure I would not have been happy doing such work. It is true that I spend quite a lot of time at work daily, but I am generally content with how intellectually stimulated most of the time.

I do like to multi-task. This means that I like to carry on several projects the same time. This, of course, results in a heterogenous efficiency rates for each project. However, this methodology offers me an escape once I face a struggle in each project. I find it very helpful to immediately switch to a different task when I am stuck on one. Someone might say that this would create a lot of confusion and it would decrease effectiveness. This is possible at times of course, but generally it has been working well for me.

I expect myself to be the main driver of my project. This being said, I expect my supervisors to be reasonably available when I get stuck in areas where I lack experience. It is more important for me that the supervisors help me identify areas of knowledge and learning outcomes rather than to direct to specifically what I must do. I already receive a lot of supervision on weekly basis that is driven by my needs. I have not faced any problems this far. My supervisors may not always be experts in specific techniques I would like to pursue, thus my expectation in this case would be that the supervisors help establish new collaborations with persons who are able to teach and supervise me in such occasions. I luckily work in a multi-disciplinary environment where additional help is easily accessible.

I generally prefer to handle everything on my own. This of course has many interpretations and implications. As I said earlier, I prefer to be the main driver of my project. But of course, I will not be able to accomplish any of my work without the help of my supervisors. I am sure that there will be a lot of administrative hurdles that I would not be able to deal with myself. A discussion point in the intro to PhD course was the writing of a manuscript. I realized people have widely different opinions on how it should be done. In my opinion, if I am the first author of a paper, I should be responsible of the way the paper is written. Of course, I may lack the experience to produce a good manuscript. The role of my supervisor should be to direct me to produce a good one rather than to do it for me.

On the other hand, I mostly fear to manage the administrative hurdles. Part of my goals during the PhD is to learn how to manage administrative tasks and so. But I can foresee difficulties in that regard. I am certain that my supervisors will be able to help me overcome any hurdles.

The course requirements for the PhD program add up to 26,5 hp. The compulsory courses are: Introductory course, Oral communication, Research Ethics, Theory and Practice of Scientific Communication, Knowledge in collaboration - how free research creates values in society, Applied Qualitative Methodology, Applied Statistics I & II, Laboratory Animal Science. Comprehensive Generic Skills (Portfolio), and One elective course. So far, I have followed the Introductory course and Laboratory Animal Science.

As for the extra course work, I plan to take as many of the teaching non-compulsory courses such as problem-based learning. I intend to continue in an academic career, thus it is important for me to take these courses when I can.

I do think internationalization is very important. In fact, I am not from here and Sweden is the 5th country I live in to pursue my academic career. I can say that without this international experience, I would have not been the same person at all. Overall, experiences are what make us who were. So, the same goes for the PhD program, internationalization in this case would entail going abroad to different labs to learn and share knowledge across. I do plan to take parts of my PhD to go to different labs in different countries to achieve different goals.

I believe that my choices this far have been aligned with my future goals of pursuing a career as an academic researcher. But this is only the beginning. Anything could happen. But I am positive that I am on the right track for my career.

We have faced several challenges in the building of a new lab when we moved to Sweden, but it feels great to have gotten this far. Our older location at BMC C12 was not sufficient to house all our needs for experimental space and equipment. Luckily, we have found a new location at BMC B10. Now that we are moving, it feels like a fresh beginning.

This time we have more space and we already have most of what we need. It is very interesting to have to participate in the thought process of where things should go. Every piece is placed where it is for a reason, so it can be utilized for the best.

This is a unique learning opportunity as most PhD students do not get to contribute to the environment they perform their research.

It is a privilege to have the opportunity to apply for grants to join international conferences. I think this experience is very important for pursuing the research process. Later in my career, I will be applying a lot more for grants.

I have been accepted to present a poster at the Keystone Conference on Signaling dynamics and Cellular plasticity in Keystone, Colorado, USA. This conference is great for me as it is multidisciplinary and there is a lot to learn from all the fields and to apply in my field.

I applied for the Fysiografen grant and I was awarded 16 000 SEK to cover some of the conference costs.

Update: 2019-09-01 To complete the process of handling grants, I have submitted a grant report that describes my activities. Download Here!

Update: 2022-05-01 I have applied to the fysiografen research grants twice after: Once rejected and later another accepted. See reflection in this section. See also reflection on the rejection in the Career Developement Section.

My halftime was done on the 28th of October 2020. This was at the beginning of the period in which the university started implementing distancing rules and limiting the number of people that can be in a room.

We held the half time at Dora Jakobson in BMC D15 and the only people present were the opponents and PhD supervisors. Everyone else was invited to join on Zoom. While this aspect was new to us, it was also a great opportunity for me to have my family (from Gaza) join us in the half time.

Choice of opponents. My opponents was Dr. Johan Jakobsson and Dr. Filipe Pereira, both of which are outside the lung field. The choice of the opponents was strategic for several reasons. First, the expertise of the opponents is very important to advance my PhD studies; Dr. Jakobsson has a great expertise in stem cells and chromatin modifications which is perfect for me since I have started working with Cleavage under target and release under nuclease (CUT&RUN), and Dr. Pereira is an expert in cell reprogramming and transcription factors role in stem cells and cell reprogramming. My focus of the PhD is on identifying context-relevant transcription factors in the stem cells of the lung epithelium that my play a role in fibrosis.

The aftermath I felt very good during the presentation and managed to comfortably explain all the different aspects of my PhD studies. (list of planned papers at the time). I was able to have a good conversation with my opponents that was focused on the bigger picture an the relevance of my project, rather than experimental details of the different experiments. To me, this indicates that the quality of the experiments was high.

I hope that I will be able to complete my PhD despite the spread of the corona virus. While Sweden is not shutting down completely like other countries, we are already seeing changes in the dynamics of the different facilities and labs at the university.

I have so far applied to three grants to the Royal Physiographic Society (Fysiografen). I was lucky to receive the first one which was a travel grant with amount 16 000SEK. The second one was a research grant that was rejected. The third was another research grant that was accepted luckily and I received 50 000SEK towards my research project.

There are a few similarities and differences between the two research grants. Here are the most important aspect that I think contributed to the decision of each grant:

• Timing. I applied for the rejected grant before my halftime review, and the one accepted was one year after the halftime. The fysiografen states that they give priority to PhD students who complete their halftime. While I was aware of this, I decided to apply anyways in order to practice research grant writing. Unfortunately, there weren’t any feedback given with the rejection.
• Amount. The first grant that was rejected was supposed to go towards the purchase of a plate reader that potentially was going to cost 400 tSEK. The second grant was to cover the costs associated with mass spectrometry (MS) proteomics and the application totaled to 100 tSEK approximately. The grant call does not specify the amount to apply for, however, I can imagine that the first grant was over what the fysiografen usually awards PhD students. I have observed after being awarded the second grant that the fysiografen usually awards 50 tSEK to PhD students and 80 tSEK to PostDoc. Therefore, I think the fact that the first application was very high in amount, it may have discouraged the reviewers to consider it.
• Experiments vs. equipment. It seems to me that this type of grant is more suitable for research projects that are specific to experiments rather than equipment. While the design of the project for the first (rejected) grant included specific experiments using the specialized plate reader, the costs associated with that project was to planned to cover the costs of the device. Since this is a smaller grant, the fact that the granting agency gives less money may make it more difficult for them to consider a grant application for an individual piece of equipment that may have a large cost.

Overall, I think this was a positive experience in general and I do think that my grant writing skills have improved through the PhD. This improvement is a result of grant applications and also helping my supervisor with larger grant applications.

This Year I am going to have three bachelor students simoltanously and will co-supervise 1 master student.

Out of the three bachelor students, two are from the Universidad Francisco de Vitoria in Spain: Laura and Pablo. See teaching section below.. And the third student Pieter is from Balgium. For Laura and Pablo, I needed to provide their University and Erasmus with a project plan before they come for their internship. For pieter on the other hand, the project plan was to be done after his arrival.

I have done this previously with Eliora who interned with me for 10 weeks in the summer of 2019. For Eliora, we have done a back and forth feedback to generate the project plan. This process was very useful for both of us as I managed to explain all the necessary information for her and easily identify the gaps in knowledge to be able to come up with the plan. On the other hand for Laura and Pablo, due to the time limitation, I have firs designed their project plan with the intention to go through it on more time with them when they are here. This way, they can apply for erasmus scholarship and also get to learn how to improve project plans afterwards.

Both of these projects were spun out of my PhD project but were made specific enough so the students get to generate new novel data and experience the research process from beginning to end.

Download: Laura’s Project Plan, Pablo’s Project Plan

I just received an email with the approval letter for my most recent application to use the SNIC local storage cluster.

This is my 6th application to be approved in association with LUNARC computation cluster.

This resource, especially, has been very instrumental throughout my PhD and is very important for anyone doing computation. We are lucky to have access to such resources at no extra cost. The only thing we need to do, is apply.

The application process for this is very comprehensive as it requires you to explain how the computation cluster is needed for your specific project. More importantly, you apply as the PI which is of course more responsibility, but a great learning opportunity.

Examples of application and report submitted after completion of project:

Application: LU local compute
Activity report after completion

In order to be able to manage the bioinformatics analysis during my PhD. I took the approach of learning how to do the different bioinformatics pipelines on my own. I have had programming experience previously, but I only started doing data analysis during the PhD. I taught myself how to use the statistical software R through online courses, guidelines, and practice. Following that, I took some advanced courses with NBIS.

NBIS has been a great source of training for me where I have learned a lot about bioinformatics analysis. I think I have developed my skills a lot with the courses I have taken there:

SciLife Labs and NBIS: Omics Integration and Systems Biology SciLifeLab/NBIS, Stockholm, Sweden

RaukR 2018. Advanced R programming for Bioinformatics Summer School SciLifeLab/NBIS, Visby, Götland, Sweden

In addition to these course, doing work with own datasets is maybe the most rewarding and best approch to learn bioinformatics.

Now at the end of the PhD, I have developed several pipelines for RNAseq, single cell RNAseq, DNAseq, CUT&RUN, ChIPseq, TAXONOMY, and more. I think I have a huge appreciation for bioinformatics, artificial intellegence, and machine learning that I plan to do choose career path that heavily uses bioinformatics.

I do enjoy programming a lot. This is why I have designed this page from scratch to display my PhD protfolio.

Dr. Richard Ingemansson is the head of thoracic surgery department and one of the surgeons performing the lung transplantation.

I had the opportunity to show Dr. Ingemansson our work with the Precision-cut lung slices (PCLS) and how we utilize the use of lung tissue in our experimentation. Dr. Ingemansson has been a great influence on our work at LBR and his knowledge and experience in lung transplantation is of great importance to us.

Dr. Sandra Linstedt is one of the thoracic surgeons at the hospital who performs lung transplantations on a regular basis. Dr. Linstedt is our WCMM partner and she is one of my PhD Co-supervisors. I have had many great discussion with her about the procedure of lung transplantation and about the tissues we obtain from the donors and patients. Dr. Linstedt will be a great source of information for me during my PhD studies.

I have had some experience previously with primary cell isolation from mouse and human. However, there was a substantial need to learn more to achieve proper isolation from the various types of lung epithelial tissues.

Dr. Oskar Hallgren - HBEC isolation Dr. Hallgren is a senior scientist who has joined our WCMM partner Dr. Sandra Lindstedt. Dr. Hallgren has been very instrumental in helping us set up the isolation protocols for the human bronchial epithelial cells (HBECs). Dr. Hallgren has been working in this field for a long time and it is a privilege to have someone to seek their advice.

For primary mouse cell isolation. I needed to improve the current technology in order to be able to isolate primary proximal and distal lung epithelial cells from the same individual mouse. I managed to make a methodological advance using 3D-printing and generated a surgical guide that aids this technique. This has become one of my first author manuscripts and is currently under review. Check out the BioRxiv

After having taken a few courses in bioinformatics analysis. I have been learning how to analysis single cell datasets on my own. There is a lot of information in published dataset that can be obtained by performing few analyses.

Dr. Leigh is the newest WCMM fellow who joined Lund University recently to start his own lab. Dr. Leigh explores mechanisms of limb regeneration in the Axolotl. I met Dr. Leigh the first time at the Keystone conference in 2019 when he was a PostDoc in Harvard. It is great to have him around.

Dr. Leigh has been helping me a lot in navigating single cell datasets and also bulk RNAseq datasets. Having fresh eyes from a different field is very helpful.

Moreover, Dr. Leigh helped me get started with Deconvolution of bulk RNAseq using reference single cell RNAseq datasets.

Dr. Soneji is PI at the Stem cell center. His team is developing a new tool that allows for interpretation of scRNAseq data analysis using virtual reality. I tried it today, and it is super cool. I think this technology will be very useful for all scientists.

Dr. Soneji has also been very helpful to me while I have been learning how to use the high powered computing cluster. I have had great conversations with him about optimization of my code to run on the cluster.

Dr. Cantù is a Wallenberg Molecular Medicine Fellow at Linköping University. Dr. Cantù is a developmental biologist who specializes in understanding how gene regulatory mechanisms control cell fate and organ formation during embryogenesis.

He is an expert on pull-down mass spectrometry proteomics and ChIP sequencing.

Dr. Cantù has agreed to be a co-supervisor for my studies. I am excited for the things I will learn in his lab.

Mass spectrometry (MS) proteomics is needed for my project in order to analyze the proteins that are pulled down with Yap/Taz to validate the finding from CUT&RUN.

I have started to learn the technique early on during my PhD and have done a trial run with Dr. Neserin Ali in 2019 to learn the technique.

While it is an exciting technique, I think it is more beneficial to me to acquire MS data through collaboration rather than try to the analysis myself. I think I have done enough to be able to understand the technique and the data that comes out of it. However, it will not be a technique that I want to go in depth with in the future. Thus, the best approach is to do it through collaborations.

I have been very fortunate to be exposed to several microscopy techniques during my PhD.

Imaging of 3D models is very important and we wanted to attempt at visualizing the mouse lung in 3D by imaging with fluorescence. Thus, we developed the necessary protocols to do it with light-sheet fluorescence microscopy that allows for larger sample size without destroying the sample with the lasers.

Lightsheet microscopy produces amazing large 3D images. While the amount of data is huge, it presents a challenge for data analysis. Attempting to analyze large 3D images using a normal screen in 2D has proved to be difficult.

We have looked into virtual reality to analysis 3D images. Luckily, a software has been developed by researchers to do this exact job: SyGlass. I have heard about this software through Dr. Shamit Soneji and I got in contact with the CEO of the company Michael Morehead.

Michael has been super helpful and has guided us through this technology very well. We do have the software license now and we are analyzing our data using it. Hopefully, we will be able to publish these analyses in the near future.

Eliora was my first full-time student during my PhD studies. I have trained a lot of people on the use and performance of several techniques. But this was the first time I had to train a student from scratch. Eliora is a second year bachelor student from the University Of Toronto (UofT) in Canada. She was selected to join a summer program at Lund University, we were content to host her at our lab.

I trained her on several techniques such as immunoblotting, qPCR, and Luciferase reporter assays. I was impressed by the speed of which she had learned and her ability to digest new knowledge.

I spun out a small project for her from my PhD project. Eliora explored the role of TGF-beta treatment on the activity of the co-transcriptional activators Yap/Taz in lung epithelial cells.

During her examination, the external examiners were impressed by her work. I felt so proud.

I hope that I have done a good job as a supervisor, and hope to do the same with future students too.

I have trained several of my colleagues from LBR and other labs on some laboratory techniques to aid them in their research.

Selected Examples

I have been fortunate to be able to join two courses by the national bioinformatics infrastructure in Sweden (NBIS):

2019 - SciLife Labs and NBIS: Omics Integration and Systems Biology. SciLifeLab/NBIS, Stockholm, Sweden

2018 - RaukR 2018. Advanced R programming for Bioinformatics Summer School. SciLifeLab/NBIS, Visby, Götland, Sweden

I have used my experience and knowledge learned from these courses to get my colleagues started on using R. So far I have helped visiting PhD student Woijtek and LBR postDoc John Stegmayr.

We as a group, have organized a workshop for the WCMM school on 3D bio printing.

Martina, a PhD student from LBR, has prepared a presentation for the student’s on bioprinting. I have given them a practical workshop on how to make a 3D model and showed them how to print that using our modified MakerBot.

Although, decellularization is not the main technique in my PhD studies. Being in the LBR, I have learned it early on, and I will utilize it in the later parts of my PhD.

In the meanwhile, I have helped in running the WCMM research school workshop on decellularization.

After having worked on the procurement of the cytation 5. I took on the responsibility of taking care of the device initially and make sure that everyone who is using it is trained. The device is such a complex machinery that is capable of many function. I would hate for it to get damaged for improper use.

So far, I have given the basic training for several of my colleagues at the lab and students who are joining the laboratory briefly.

While this task of device care and maintenance will be passed on to someone else in the future. This experience has taught me a lot.

Pieter has joined us from Belgium at the end of his bachelor degree to do his traineeship. His program was for him to become a laboratory assistant/Manager.

I designed Pieter’s project for him to help me with the optimization of subcellular fractionation from lung epithelial cells while ensuring mass spectrometry compatibility of the resulting sample.

Pieter presented his thesis at his home country where he received a high grade.

Title of Pieter’s thesis: Optimization of subcellular fractionation of the lung epithelial cells as exploration of the Hippo signaling pathway.

I think the amount of knowledge Pieter has gained through his internship is reflected in his thesis.

Download Pieter’s Thesis

I have co-supervised Thomas Syski who did his MSc thesis with us. Thomas’s project was titled: 3D bioprinting of alginates into structures resembling human airways

I trained Thomas on 3D(Bio)Printing throughout his project.

Check out the abstract on LTH website

It was nice to train multiple students together. We received two highly motivated students from Spain. It was a great experience to work with both of them.

I trained Pablo on plasmid preparation and cell culture techniques, and I trained Laura on Bioinformatics analysis of transcription factor motifs and single cell RNAseq.

Both of these students were highly motivated and managed to produce great project reports despite the fact that this was done during the pandemic.

Laura worked mostly from her dorm room here and Pablo had to fit his schedule to make sure that he is only present in the lab when it is possible to follow all the guidelines for the pandemic.

I am proud of both of them.

Download:
Laura’s Project Plan
Laura’s Final Report

Download:
Pablo’s Project Plan
Pablo’s Final Report

The experience of training Laura and Pablo from Spain was very postitive. Their university was very content with their outcome so they have decided to send us more students. We received another two highly motivated students from Spain: Inés Mínguez and Irene Urrecho. Ines helped me with the optimization of Immunoprecipitation protocols to pulldown Yap/Taz from lung epithelial cells while maintaining mass spectrometry compatibility. Irene has worked with me on the CUT&RUN experiements and the bioinformatics analysis associated with these experiments.

Next week is their last week and they will present their work at their university later on this summer. I am sure they both will do great.

Inés Mínguez Santos

Project title: Optimization of immunoprecipitation of Yap/Taz proteins from MLE12 immortalized murine lung epithelial cells
Download Inés’s Final Report —————————-

Irene Urrecho project title: Identification of Transcription factor motifs targeted by Yap/Taz-complexes in the lung epithelium using CUT&RUN

Download Irene’s Final Report

The Miltenyi company has acquired the Blaze lightsheet microscope and they held a workshop in collaboration with Lund University BioImaging Center (LBIC) on 2021-11-18. During this workshop, I joined the event by organizing an interactive session for people to view lightsheet imaging using Virtual Reality on the Arivis Software.

This was an opportunity that presented itself suddenly. I have been setting up the Virtual Reality for our lab to explore lightsheet images we have obtained. I was going to attend this workshop as an attendee, however, I heard that the person organizing the VR part of the workshop was not available anymore. I proposed to the organizers that I can do it, and they were happy to include me in this workshop.

I also chaired some of the talks in this workshop, which also has given me some more exposure.

Title: Relevance for me, my attitudes and what I want to learn. Several factors may contribute to a successful research career and these factors are dependent on the environment where the research is conducted and the attitudes of the research. I do believe that collaboration and innovation are among these factors that contribute to a successful research career. However, I had not given enough consideration and thought to the role of entrepreneurship. Many biomedical researchers go into their perspective fields in the goal of figuring out a disease or a problem that affect humanity one way or another. Thus, entrepreneurship is not on the radar of many biomedical researcher. Personally, I think if an opportunity presents itself through my research career, I would pursue the entrepreneurship route. However, I do not have it as a direct goal of my career.

I believe that collaboration in research is extremely important. The more we learn in our fields, the more questions we have. And there isn’t enough time for anyone to explore a research question on their own. In terms of doctoral studies, I need collaborations to learn several of the skills that I need to finish my project. Currently, I have several collaborators locally at Lund university, and generally in Sweden in addition to other international collaborations. I believe that my role in these collaborations is to trust the expertise of my collaborators and give back the same level of quality to earn their trust. Having clear assigned roles in collaboration is usually very helpful to accomplish the different goals.

My current research is studying pathomechanisms of pulmonary fibrosis. Idiopathic pulmonary fibrosis (IPF) is a lethal disease with unknown causes and no available cure. The lack of effective therapies can be attributed to the gap in knowledge of disease pathomechanisms. My research project is aimed at finding new transcriptional complexes that are active during disease. Knowledge gained in my research project can aid in finding new therapies for this lethal disease.

From the introductory lecture, there was a great focus on entrepreneurship. I would like to learn more about such opportunities and how our environment here (Lund) can be helpful in that regard. Additionally, I look forward to learning how to understand and create value of my research.

After the course: Download final reflection after the course

I have been selected to represent the WCMM center at strategic meeting the KAW foundation and Wallenberg family representatives.

The goal of this meeting was for the KAW foundation to receive updates from Lund as one of its more recent centers in order to decide the path forward and strategize their focus areas for future funding.

THIS FELT LIKE A BIG DEAL. I honestly can say that this was one of the few times during my career in Sweden where I have witnessed a new level of professionalism. It is somewhat strange, but it was good exposure for me. I like being involved in strategic meetings as I think that I can voice a lot of the opinions of my peers.

I think I did great during this meeting and was able to participate in the conversations in a positive manner.

Dr. Claudio Cantù is an expert in ChIP sequencing and CUT&RUN technologies. This collaboration with Dr. Cantù has resulted in him becoming a co-supervisor on my PhD. This collaboration continues to be very important for my PhD project.

This collaboration ended up being one of the most important ones for my project. We have managed to successfully implement the CUT&RUN technology for Yap/Taz in lung epithelial cells. This is normally very difficult to do for transcription factors, and even more difficult for co-factors like in our case. We managed to reproduce this across various samples of isolated primary alveolar type II cells.

In the beginning of my PhD, I have co-organized a symposium on the use of precision cut lung slices (PCLS) in a collaboration between Lund University and Nottingham University.

During this symposium, we have started a collaboration with Dr. Amanda Tatler to join forces on the topic. We are currently preparing a review article as part of this collaboration.

I just got invited directly to review a paper from a very respected lung journal :)

American Journal of Respiratory and Critical Care Medicine

Dear Mr. Alsafadi:

The manuscript entitled “A Highly Phenotyped Open Access Repository of Alpha-1 Antitrypsin Deficiency Pluripotent Stem Cells” has been submitted to the American Journal of Respiratory and Critical Care Medicine and I am writing to invite you to serve as a reviewer. The information of the manuscript is attached at the end of this letter.

I am very excited to review this paper, but also a little nervous, I do not want to do any mistake. I attached my review of this article.

Download Review

My main supervisor, Darcy Wagner, has trained me to do peer-review of scientific articles through doing. Every time she has gotten a new paper to review that is related to my work, she would inform the editor of that journal and let me help her with that review. We would work on the peer review together by first analysis the paper separately, second intensely discuss all aspects of the peer, and third write the main major, minor comments.

This process has helped improve the way I think about experiment design and about how I write my own scientific work.

Darcy always informs the editors of the trainees help so that the editors can recognize the expertise and maybe one day directly as a peer review.

I have recently developed a protocol to immunolabel a whole mouse lung and process it so it can be imaged using light-sheet microscopy to visualize specific molecular features within full murine lungs. We call the method: inFLATION: infusion of Fluorescent Labelled Antibodies in Tissue of Intact OrgaNs

This technique have showed great success in wildtype lungs. I have presented this data at the EMIM conference in Greece as mentioned section 6. One of the main questions I have received about this technique was whether this protocol is able to image diseased lung where normal lung architecture is damaged.

To address this question, we have collaborated with Dr. Irma Mahmutovic Persson, who is an expert on MRI and micro CT scans of rodent lungs for detection of lung fibrosis. In this collaboration, we have imaged the lungs using MRI micro CT and followed that by processing with our protocol to compare the imaging modules and to evaluate whether our protocol is capable of properly staining the diseased lung tissue.

This collaboration has developed over time to include several aspects. We do qPCR and immunoblotting in our lab and that’s why Roberta, another PhD Student, and her PI Iben approached us for this collaboration.

This collaboration started by providing my help to Roberta and her student Linda on qPCR of the gene Aquaporin 4 (Aqp4). I have trained both of them and helped Linda throughout the whole process of Data acquisition.

The first version of the manuscript was not accepted. Thus, we decided to focus the manuscript on distinguishing between the different isotypes/variants of Aqp4. Thus, this collaboration grew further for us to design and validate new primers in addition to optimization of protein extraction from brain tissue and the immunoblotting of Aqp4.

If you ask me before going through this, how long it would take? I would have said it is very quick and painless. But the opposite has proved to be the case. Dealing with Aqp4 was very tricky and this collaboration took very long.

Luckily, we were able to pull it off and optimize all the technical aspects for this study and the manuscript is now submitted again to a different journal.

Membership in Scientific Societies

My research explores the role of specific co-transcriptional activators (Yap/Taz) in the lung epithelium in the context of Idiopathic Pulmonary Fibrosis (IPF). IPF is a lethal chronic lung disease with no cure. The exact causes of the disease are unknown and median survival is 2.5 to 3.5 years. Current available therapies are only able to slow down the disease progression. To date, there isn’t a therapy that can stop or reverse this disease. The molecular mechanisms of disease progression involve several signaling pathways in the different cell types of the lung. The specific role of the lung epithelium is not well understood. The aim of my research project is to identify new transcriptional complexes that are relevant to IPF pathogenesis.

The approaches of my research studies are highly reliant on the use of primary cells. We thus isolate primary epithelial cells from both mouse and human tissue. Of course, there are ethical considerations for the use of animal or human subjects.

In the mouse experiments, the aim to induce fibrotic characteristics using the administration of Bleomycin. This model of fibrosis is well established and commonly used. We have obtained the ethical approvals for appropriate durations of the length of the experiments. There are not many ethical issues with the cell isolation after the animal has been sacrificed. However, the use of the specific cells in our studies will help answer our research question directly. Thus, this contributes to the concept of the 3R as we will not need to sacrifice many mice for the purpose of the study. A big part of my work is to optimize these isolation protocols to cut down the number of animals used to less than half of what is equivalently used in literature.

As for the use of human tissue, we have a partnership with the thoracic surgery department at the hospital. We have been granted the ethical approval to use excess tissue from transplant surgeries and possibly failed transplants. These tissues would be disposed of otherwise. Thus, we have obtained the ethical approval to use these tissues and all the necessary considerations are taken into place, such as informed consent. The way it is done in Sweden is that the person receiving a donated organ is the responsible person for what happens to that tissue. We thus get informed consent to use this tissue from the recipient of the donor lungs. Of course, all these procedures are taken care of at the hospital. At our preclinical lab, we only get the tissue if it has been approved to be sent to us. I mostly do epithelial cell isolations from these tissues. Thus, one ethical consideration that is always taken in consideration is the privacy and the dealing with sensitive information. It is of course highly unlikely to identify someone solely by the tissue, but we anyways have taken a lot of measures to avoid any breech of this privacy. When we receive a human tissue material, we assign a new identifier for that piece of tissue and all data regarding the patients/donors are not provided for us. The only information we obtain are the age and gender of the of tissue as approved. The reason we need this information is because they play a major role in the behavior of the cells obtained from these tissues (i.e. cells from older tissues do not have the same proliferative capacity).

We are highly privileged to be able to perform studies on primary tissues. We are always grateful for that and we do treat these tissues with the most respect.

Andréa Wiszmeg from the Department of Social Work at Malmö University has reached out to conduct an interview with me with regards to a project on ethics they are running.

Among the aims of this project was to undertand how “responsible researchers” are fostered. Here is a quote from the abstrac for their project:

One of the aims of the project is to understand how “responsible researchers” are fostered. We look into how ethical reflexivity is expressed, practiced and understood in the day-to-day of biomedical research environments. But what happens to our knowledge production when we make biomedical researchers engage in this bird’s-eye view on their research and its socio-cultural circumstances? How can neither taking a distanced position, nor engaging in direct activism but rather pushing toward areas we consider possible hotbeds for public debate be further theoretically conceptualized? What does such an endeavor imply for our role as STS scholars? And how does that connect to the tendency to engage humanities and social science scholars as interpreters and mediators in cross-disciplinary projects in knowledge-intensive policy fields, such as ours ? In what way is the symmetry principle (Bloor, 1976) affected, when value-laden initiatives as ethical reflexivity and public engagement is treated as an inherent good or as “truths” to be pursued?
Source

In this interview, we have talked about the use of 3D bioprinting in research and what are the ethical considerations that we take in order to maintain our reponsible research behaviour. We have also talked about the use of primary cells in research and the ethical aspects of that.

There is no doubt that the big data has changed the way we do science and have opened so many new avenues that allow us to explore our data better. There are many techniques right now that utilize big data or produce big data such genomics, transcriptomics, proteomics, metabolomics, and many other omics. However, there is an important issue that is associated with this. Which is when you produce big data from a patient sample, it is as if you are creating a fingerprint or signature of that person’s molecular makeup. But the level of information you can obtain from each element is variable of course. And it is not always clear whether you get identifying information from that comic or not. So maybe one day. You imagine that your big data does not have any identifying information about your patients, however. Somebody can at some point produce a technique, or a methodology that will obtain such data from your data set, since our bioinformatics pipelines are increasingly improving and there’s a lot more information that we can learn from data by simply doing some mathematical models on it. Therefore, the question becomes, how do we deal with this when we are dealing with patient samples? Of course, the ethical committees that approve studies that contain big data have thought about this and it is becoming a requirement for people. To include how they will save and secure the patient data. This is certainly an interesting topic. While patient information must be secured at any cost, there should always be enough access to the data used in any analysis. To be able to fully evaluate the science that is produced. I think that a lot of the main institutes are trying hard to address this issue and there are many regulations that are associated with it. And there are many big institutes that provide Tools such as NCBI or ENI. However. These tools are not usually available for all types of data. An example, there are several recent advances in imaging that allow us to produce high content high quality images of samples that may one day be able to contain identifying information that can identify the patient of that sample. Such as this specific marker. Or things that are associated with disease. In my own research, I usually do my best to deidentify all the samples that I have. And make sure that there is no information that can be used to identify the samples that we are using. I think the field is moving in a good direction and that there is a lot of effort going into preventing problems in this regard. However, I think it is important for all of us who are working with this type of data to be careful with this and to be considerate of the patient material that we get.

In reading My Portfolio so far, it is probably noticeable that I talk a lot about big data and the type of things we need to analyze it and store it. A very important resource that has been available during the PhD was the LUNARC cluster. High-powered computation is very important for this type of analysis because you require a lot of RAM and a lot of CPU in order to be handle such large datasets. However. Do you have computing systems need to be maintained? And there are of course many ways to do that, one of which is to be able to update these systems online. So, in many cases, the high computing cluster is connected to the Internet, so the software packages can be updating themselves automatically or manually. So, when having any system connected to the Internet. This system becomes susceptible to attack. And that it’s possible that any type of data can be stolen from that system. Our high computing cluster is quite secure and requires multifactor authentication to be accessed. However. It is still important to be able to secure data and especially patient data. Luckily. We have the LUBI cluster. LUBI Is another computer cluster That is managed by the stem cell center and is used for secure data. This system is disconnected from the Internet and can only be accessed while you are present at the BMC. I think it’s great that we have access to the system Even though it requires a lot of more work in order to be able to analyze data on it. The reason is that you require a lot of software to be installed manually on another machine and then transfer it to install it on the secure cluster. Of course, it is worth it to do all of this to be protective of patient data.

Currently, there isn’t a similar system to use for analyzing imaging data. Therefore, it’s important to try to fight for resources to be able to create such a system.

I have been working in the research field since 2015 which is not a long time, but fairly long enough to learn about my learning abilities. I have realized that I am very particular about how I learn things, and more importantly how I like to be supervised. I consider myself very independent and I always like to try to solve issues on my own before pursing the answer from some one else.

I am lucky to have a unique relationship with my supervisor. We do discuss the supervision styles quite often, not only how she supervises me, but also how I can supervise others. It is a chain after all.

LBR lab, an example, consists of more than 15 members, each of which has a unique way of learning. It is important to always observe how people perceive information and absorb new content.

I started the PhD program knowing that I will have to reflect on supervision as the project progresses.

We were quickly able to identify that we needed a supervisor with strong biochemistry expertise in order to address the transcription factor identification part of the project. My main supervisor, Darcy Wagner, has met Dr. Cantù in one of the WCMM retreats. I contacted him and scheduled a skype meeting. I described my project to Dr. Cantù and we immediately agreed that he can be very instrumental in my PhD studies.

A few months later, we planned a pilot study and that was a confirmation for both of us that he would not only be a collaborator, but also a supervisor.

I am glad to have found Dr. Cantù and I look forward to the rest of my PhD.

I was fortunate to be in the LBR lab before starting the PhD. I was able to understand the capabilities and limitations of myself and my environment. My PhD project was designed by myself and my supervisor. We worked together on finding the best experiment designs to address all questions raised under the unified aim of the study.

The process of study design was done in tandem with writing a VR starting grant. Darcy wrote her starting grant on this project. We sat through several meetings and brain-storming to reach the final design of the project. I helped Darcy with the VR grant mainly in experiment design and figures. This experience was very useful for me to think at a higher level of project design and also to learn more skills in grant writing and creation.

I am thankful to have had that opportunity to take a project from its very beginnings and I hope to see it through to a good place by the end of my PhD.

Adapted from section 1. See reflection in section 1

The aftermath I felt very good during the presentation and managed to comfortably explain all the different aspects of my PhD studies. (list of planned papers at the time). I was able to have a good conversation with my opponents that was focused on the bigger picture an the relevance of my project, rather than experimental details of the different experiments. To me, this indicates that the quality of the experiments was high.

I hope that I will be able to complete my PhD despite the spread of the corona virus. While Sweden is not shutting down completely like other countries, we are already seeing changes in the dynamics of the different facilities and labs at the university.

The pandemic has affected us one way or another. While the university has not shut down completely during the pandemic, it was evident that there are several changes that affected everybody. For example, there were a lot of issues with ordering, a lot of delays with receiving shipments and materials to use. There was a lot of shortage in materials needed for protection, such as gloves. Moreover, it was not clear. Where there will be extensions for PhD students due to the pandemic or not. That definitely has affected the progress of the PhD. That also has affected the way the supervisor has to deal with this. I think it is a two way problem. The supervisor has to implement a new method of supervision and the supervisor has to also communicate differently about what they’re going through. I think overall. We managed very well to deal with the pandemic in my lab. Here are some examples of the adjustments:
Less IRL meetings.
The availability of systems that allow online meetings has been a great addition to our routines. it is important to know that there is a way to communicate even though you cannot meet in person. of course, it took us a while to be able to adjust to this method of communication however I think after a while it was very useful and I think even after the pandemic we are still using any online systems on daily basis that fill the gap in communication. But this of course meant that you have to actively think about scheduling meetings with your supervisor in order to be have access to them. While it is a great behavior to schedule meetings before hand, it is sometimes difficult to Evaluate whether you need that help early in advance.

Less experiments vs. different types of experiments.
The delays in receiving materials and closures caused by the pandemic have forced us to change our daily routines and focus on tasks that require computer work more than experiments. But this does not mean that we are not able to do experiments. On the contrary I think this gave us an opportunity to evaluate how much experimentation we need versus how much time we need to think about the experiments we do and the time we need to analyze it. An example, there are sometimes experiments that you shelf to look at later and it takes a long time before you can analyze them. I think the pandemic has helped us to analyze a lot of these experiments, and at the same time has taught us to be able to evaluate whether we need an experiment. I was lucky personally because I just had generated an RNA seq data set. which meant that I could spend a good amount of time analyzing it. this was a great experience for me, and it has deepened my love for bioinformatics and I think I will implement more of the bioinformatics in my future career.

Supervising students during the pandemic. I’ve had several students during the pandemic some of which we’re able to perform experiments in the lab and others were able to do their products online without having to come to the lab. while there were some struggles, I think we managed to produce great work from these students. you can see some of the reports in the teaching section of this portfolio. I can say that I enjoy supervising students while they are here in person, however being able to teach them while they are online gives me more time to focus on my own study and also gives them their time to be able to develop on their own. I think it was great for them to experience that and I think they will benefit from it in their future.

Effect on career goals.
As I have mentioned earlier, I focused more on bioinformatics during the pandemic. I found it to be very interesting and very exciting for me and I may decide to transition into bioinformatics fully after the end to PhD.