User:Z3336156

From CellBiology

--Z3336156 15:13, 8 March 2012 (EST)

Lab Attendance

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Lab 1 (week2)

http://www.jove.com/basic JOVE Lecture 2

Cell Nucleus

Hi guys!

Lab 2 (week3)

Collapsing Actin Mesh.jpg

homework:

The exact location of nucleiods which contain mitochondrial DNA has been vaguely described and was a mystery due to the lack of available technology, which could measure precise locations of minute organelles molecules, that lie inside organelles. However, since the discovery of superresolution microscopy scientists were able to study further the unsolved questions of nucleiods found in the mitochondria. Brown et. al were able to discover the shape, size and relative locations of nucleoids with superb imaging. Three dimensional images showed that mitochondrial DNA are found in ellipsoidal nucleoids and is arranged in an extremely condensed fashion.


Reference: Timothy A Brown, Ariana N Tkachuk, Gleb Shtengel, Benjamin G Kopek, Daniel F Bogenhagen, Harald F Hess, David A Clayton Superresolution fluorescence imaging of mitochondrial nucleoids reveals their spatial range, limits, and membrane interaction. Mol. Cell. Biol.: 2011, 31(24);4994-5010 PubMed 22006021


--Mark Hill 12:50, 20 March 2012 (EST) Good explanation of the study findings.

Lab 3 (week4)

Homework:

Journal Articles About VEGF:

1) The journal article was researched about the effect of Hydrogen Sulfide to VEGF during myocardial infarction. VEGF binds to the tyrosine kinase and fms-like tyrosine kinase receptor that can up-regulate or down regulate vascular organisation. The research showed that mice treated with hydrogen sulfide after an induced myocardial infarct had higher levels of VEGF and the 2 receptors mentioned earlier that resulted in a better recovery after the injury.

Reference: Natia Qipshidze, Naira Metreveli, Paras K Mishra, David Lominadze, Suresh C Tyagi Hydrogen sulfide mitigates cardiac remodeling during myocardial infarction via improvement of angiogenesis. Int. J. Biol. Sci.: 2012, 8(4);430-41 PubMed 22419888


2) In the research conducted by Tororovic et al. they studied the correlation of the presence of VEFG and the proliferation of leukemic blast cells and microvessel density found in acute lymphoblastic leukemia(ALL) patients. The study found that increased VEGF leads to higher levels of leukemic blast cells and microvessel density count. This finding can be used to seek new methods in combating ALL.

Reference: Milena Todorovic, Ziv Radisavljevic, Bela Balint, Bosko Andjelic, Vera Todorovic, Maja Perunicic Jovanovic, Biljana Mihaljevic Increased angiogenesis-associated poor outcome in acute lymphoblastic leukemia: a single center study. Appl. Immunohistochem. Mol. Morphol.: 2012, 20(5);488-93 PubMed 22417860


3) The article talks about the effect of VEGF which binds on VEGFR-1(vascular endothelial growth factor-1) and VEGFR-2(vascular endothelial growth factor 2) that affects the astrocytes in terms of gap junctional intercellular communication, cell proliferation and motility of astrocytes. The experiment was conducted through the nurturing of astrocytes in VEFG rich solutions, from here, it was discovered that VEGF promotes gap junctional intercellular communication, increases cell mitosis and enhances the motility of astrocytes through the VEGFR-2.

Reference: Ricarda Wuestefeld, Jingchen Chen, Karl Meller, Beate Brand-Saberi, Carsten Theiss Impact of vegf on astrocytes: analysis of gap junctional intercellular communication, proliferation, and motility. Glia: 2012, 60(6);936-47 PubMed 22431192


4) VEGF plays major role in the progression of osteoarthritis thought the proliferation of blood vessels in the subchondral growth plate. It was hypothesized that VEGF plays a role in the early stage of osteoarthritis as a marker for diagnosis that can be used as a marker for diagnosis and was proven to be correct.

Reference: H Jansen, R H Meffert, F Birkenfeld, W Petersen, T Pufe Detection of vascular endothelial growth factor (VEGF) in moderate osteoarthritis in a rabbit model. Ann. Anat.: 2012, 194(5);452-6 PubMed 22429866


SDS for Methanol:

Properties: state: liquid appearance: colourless odour: alcohol-like, weak odour solubility in water: miscible

Hazards: POISON. Ingestion can be fatal and cause colour blindness. Vapour is flammable and still harmful. Harmful if swallowed, inhaled, or absorbed through the skin. Eye, skin, and respiratory tract irritant.

Potential Health Effects Eye: May cause painful sensitization to light. Methanol is a mild to moderate eye irritant. Inhalation, ingestion or skin absorption of methanol can cause significant disturbance in vision, including blindness.

Skin: Causes moderate skin irritation. May be absorbed through the skin in harmful amounts. Prolonged and or repeated contact may cause defatting of skin and dermatitis. Methanol can be absorbed through the skin, producing systemic effects that include visual disturbances.

Ingestion: May be fatal or cause blindness if swallowed. Aspiration hazard. Cannot be made nonpoisonous. May cause gastrointestinal irritation with nausea, vomiting and diarrhea. May cause systematic toxicity with acidosis. May cause central nervous system depression, characterized by excitement, followed by headache, dizziness, drowsiness, and nausea. Advanced stages may cause collapse, unconsciousness,

MSDS for METHANOL

Lab 4 (week5)

Musashi Protein Nicole A Siddall, Eileen A McLaughlin, Neisha L Marriner, Gary R Hime The RNA-binding protein Musashi is required intrinsically to maintain stem cell identity. Proc. Natl. Acad. Sci. U.S.A.: 2006, 103(22);8402-7 PubMed 16717192

Musashi is responsible for the balance between germ-line stem cell renewal and differentiation. Its function in stem cells is for maintenance of stem cell identity. The paper also found that Musashi plays various roles in the different stages of germ cell differentiation in the male meiosis(spermatogenesis).


Primary Musashi Antibody: Musashi-1 RBT REC OLIGO AB

Musashi-1 ABfinity™ Musashi-1 SDS for RBT REC OLIGO AB

Antibody:Musashi-1 RBT REC OLIGO AB

Host: Rabbits

Price: $280.00 (usd) /100 µG

Clonality: polyclonal (oligoclonal)


Secondary Musashi Antibody:

Anti-antibody: Alexa Fluor® 488 Goat Anti-Rabbit IgG (H+L) *2 mg⁄mL*

Host: Goat

Reactivity: Rabbit

Lab 5 (week6)

Image Relating to Topic

Alexa Fluor® 488 Goat Anti-Rabbit IgG (H+L) *2 mg⁄mL*

MSDS for Alexa Fluor® 488 Goat Anti-Rabbit IgG (H+L) *2 mg⁄mL*

VEGF signalling pathway .gif

Reference: F J Giles The vascular endothelial growth factor (VEGF) signaling pathway: a therapeutic target in patients with hematologic malignancies. Oncologist: 2001, 6 Suppl 5;32-9 PubMed 11700390


Lab 6 (week7)

Tm4-Control Graph.png

Do you see a difference in phenotype morphology between Tm4 over –expressing cells with the control group?

Fan– the control group had almost four times the number of cell count as compared to Tm4 over-expressing cells

Broken Fan – the Tm4 over-expressing cells showed a 65% reduction in cell count in comparison with the control group

Stumped - the Tm4 over-expressing cells showed significant reduction of 30% in cell count in comparison with the control group

Pronged - the Tm4 over-expressing cells showed a great increase of 320% in cell count in comparison with the control group

Stringed - the Tm4 over-expressing cells showed an immense increase of 250% in cell count in comparison with the control group

Pygnotic - the Tm4 over-expressing cells was only 17% of the total number of control group cell count

Comparison of Tm4 over-expression and the control group

Tm4 Group:

-most of Tm4-overexpressers had a stringed or pronged phenotype

-there were few stumped and broken fans and very less fan

-there were just over a couple of cells with pygnotic phenotype

-the cells in this group were brighter and more fluorescent

-cells had more branching and longer processes


Control Group:

-most of the cells had a broken fan and stumped phenotype

-there were several fan, pronged, stringed and pygnotic cells of about the same amount

-cells had lesser branching and shorter processes

-the cells were not as fluorescent as compared to Tm4 over-expressers


How could Tm4 over-expression lead to this difference?


Tropomyson is an actin binding protein that affects the activity of actin. From the lab, it was found that Tm4 over-expression caused and increase in number of branching and length of processes in B35 cell phenotype. High levels of tropomyosin induce actin activity and prevent its disassembly. Further more, it increases the availability of actin to other actin related proteins leading to more developed filaments.[1] The differences seen from the two groups were caused by the over-expression of Tm4 in B35 neuroblastoma cells. Tm4 is found to increase the interaction of actin filaments that affected the cellular phenotype, for example, longer axons and dentries on neurons. Hence, over-expressers B35 cells had longer and more processes and branching. [2]

[1] Uno Lindberg, Clarence E Schutt, Robert D Goldman, Maria Nyåkern-Meazza, Louise Hillberg, Li-Sophie Zhao Rathje, Staffan Grenklo Tropomyosins regulate the impact of actin binding proteins on actin filaments. Adv. Exp. Med. Biol.: 2008, 644;223-31 PubMed 19209825


[2] L Had, C Faivre-Sarrailh, C Legrand, J Méry, J Brugidou, A Rabié Tropomyosin isoforms in rat neurons: the different developmental profiles and distributions of TM-4 and TMBr-3 are consistent with different functions. J. Cell. Sci.: 1994, 107 ( Pt 10);2961-73 PubMed 7876361


Lab 7

Contributions for group project:

Normal Function

Vascular Endothelial Growth Factors (VEGF) bind with the Vascular Endothelial Growth Factor Receptor (VEGFR) which initiate a cascade of signal resulting in vasculogenesis, angiogenesis and lymphangiogenesis in cells.

Several factors causes the production of VEGF namely:

Hypoxia

All cells need oxygen to survive. However, in some cases not enough oxygen is supplied to tissues from various reasons, decrease in inspired oxygen from places of high altitude and preterm birth. Hypoxic cells, cells without adequate supply of oxygen can stimulate the production of VEGF. When cells are lacking oxygen, it produces Hypoxia-Inducible Factor (HIF) a transcription factor that stimulates the release of VEGF. The free VEGF then binds to VEGFR on cell membranes of endothelial cells causing angiogenesis and vasculogenesis as a cellular response.

Oncogenes

An oncogene is a gene that can latently cause cancer. Oncogenes can be observed in high levels in tumour cells. Cell death or apoptosis is a normal process of a cell cycle, however, oncogenes suppresses this mechanism; hence, leading to further proliferation of mutated cells. Some of the manifestation of oncogenes is the error in protein structure that affects the level of enzyme activity and cell regulation. In cells with high levels of oncogenes, levels of kinase production are altered. Kinases are enzymes that add a phosphate group to different proteins which acts as an on and off switch as well as receptor kinases adding phosphate group to receptor proteins found in the surface of cell membrane to send out signal from the environment to the inside of the cell. Since VEGFRs are tyrosine kinase receptors, an alteration in the signaling pathway can cause cancer by switching on the receptor in spite having no signals from outside the cell.

Hormones

-Angiotensin II

-Thyroid Stimulating Hormone (TSH)

-Corticotropin-Releasing Hormone

---Adrenocorticotropic Hormone

-Steroids?

Other growth factors and cytokines

VEGF are also produced as a response to other growth factors and cytokines. Cox-2, cyclooxygenase-2 induces the production of VEGF. This correlation has been found with the study of tumors. PMID:11326316.

Lab 8

1. Identify a mammalian cell line in the ATCC catalogue(and add a link).

V79-4 Mammalian Cell Line

2. Identify the original tissue of origin of that cell line.

Cricetulus griseus (Chinese hamster)

3. Identify the original paper that characterised the properties of that cell line. Klaus G Steube, Anne-Leena Koelz, Hans G Drexler Identification and verification of rodent cell lines by polymerase chain reaction. Cytotechnology: 2008, 56(1);49-56 PubMed 19002841


Lab 9: Peer Review

Group1 : Testosterone

•Introduction is too short but well referenced. This can be further improved by putting more information and giving a brief overview about the topic and what the webpage will be talking about.

•History: Very well formatted and well referenced. Aren't there any other discoveries after 1994 about this topic which you can still include?

•Biosynthesis: Good subheading that other groups do not have. The step by step process made it easier for the audience to understand how testosterone is derived from cholesterol and all information presented were well referenced.

•Regulation: Information were presented in a nice and succinct manner. Maybe adding a picture of the negative feedback mechanism of testosterone regulation will be a good idea.

•Signaling pathway: Like the regulation subheading, this section was presented well. Having sub headings made the division of idea easier to understand.

•Normal Function: This showed a great research about the topic. Putting subheadings will be a good idea as having massive chunks of paragraph looks overwhelming and not very appealing to readers. and The formatting of the paragraph are different. Some paragraphs had 1.5 spacing whilst the others had paragraphs that are single spaced. (this is just minor though).

•Abnormal Function: The abnormal section would look more interesting if there were images for the diseases.

•Clinical Uses: This is a good subheading that other groups did not have. Nice addition to the project. Very easy to read.

•The abnormal section would look more interesting if there were images for the diseases.

•Current and ongoing research: Interesting researches were presented in this section and each were explained thoroughly and the language used makes it easy for readers with very little background about the topic understand the topic well.

•More than one image is taken from wikipedia. (steroidgenesis and The binding of testosterone to an androgen receptor within a cell)

•This project is almost finished. All the section showed intensive research and are explained well.

Group3 : Extrinsic Apoptosis

•Introduction: This section gives a good definition of apoptosis. However it is still unfinished and needs to be referenced.

•History: Again: the history looks good but references are needed.

•Signaling Pathway: This section needs to be worked on. Putting an image of the pathway would help a lot in making the description understandable. Trying to understand it though reading was a bit of a challenge, but for sure, if an image was added this section will be good. Also, i can see that there are references, however, they are not linked properly. I understand that you're still working on this section but don't forget to add them.

•Function: This section is still unfinished and could be improved better. It was quite hard understanding it because there were several technical terms that were not explained well.

•Current research: The referencing of this section isn't in a wiki form. You can look at other pages to see how they referenced their work. In addition, several diseases were mentioned in the current research introduction but only two were talked about, cancer and leukemia. More current researches could be added in this section.

•Glossary: this section needs to be filled out. Quite a number of technical words were presented that could be added in this section like proteases, necrosis, and cytotoxic.

•Some subheadings are missing like the abnormal function.

Group4 : Notch

•Introduction: The introduction is too short. It talked about what are the types of notch, its ligands and where they can be found but it failed to mention what is Notch and what does it do. I think giving a simple description of what Notch is and what does it do is the purpose of having an introduction. Despite of this, the section is well referenced.

•History: The table format of this section makes it easy to read. I think this section could be researched more. I tried searching Notch on pubmed and hundreds of journal articles were found, therefore, there are more history that you can put in this section.

•Pathway: This section is too short. Very little information can be read and it lacks depth. Having an image would help your readers follow what you are discussing.

•Protein and Receptors: This section looks good although not referenced. I suggest for the part "See video under external link for further understanding." You can already link the video here rather than having to scroll all the way down to watch the video. Also, there are no references given in this section.

•Normal Function: This is the best section of your project. It is well referenced and gives a simple and easy to read understanding about the topic.

•External Link: It's good to have this at the bottom but also link it will be better if it was linked to the section that it relates to.

•Further research: The further research section showed little research. Only titles of articles were given and their corresponding links. This could be improved by giving a brief description of each current research linked.

Group5 : Wnt-Beta catenin

•Introduction: Introduction of this page is good. It gives just a touch of information about Wnt-Beta catenin making you want to read more about it.

•History: This section shows detailed research the group has done. It gives an intensive timeline that is well referenced.

•Mechanism of Action: This section is good, however, the flow of idea is quite hard to follow. I think it will be easier to understand if it was in paragraph form rather than in bullet point format. It's nice that a group has already posted a student drawn image.

•Disease section: This section is well referenced and the the group talked how the signaling pathway results to abnormal function. Also, the layout makes it easy for the readers to understand this section.

•Key players in Wnt/b-catenin signaling: This subheading shows intensive and well constructed research about the topic. It was well referenced and the images make this section engaging.

•Embryonic Development: This section is informative yet I think this section can be placed under Mechanism of Action.

•Future directions: There were several bullet points under this section that were not referenced. Maybe, adding articles that shows experiments being conducted in search for the answers would be good.

Group6 : Insulin

•Introduction: The section showed a good overview of what is insulin, what it does, how does it lower blood sugar level and what happens when this pathway is disturbed although there is only 1 reference and i think the subheadings are unnecessary.

•Structure of Insulin: The image is not referenced properly.

•History: This section seems need further research. Only some sections had a proper reference. Also, this can be improved by adding what PI3K, IRS-1, and GSK3 (I know definitions were given towards the end at the glossary section but it wouldn't harm telling the audience what these proteins do.

•Signaling Pathway: Well written section but i think you can let go of the 'introduction' subheading.

•Normal Function: There are far too little information given in this section. It can be improved by putting more depth and having more references in this section.

•Abnormal Function: Having subheadings in this section made it easier to read. This can be improved more by linking how the signaling pathway can cause these diseases like how the group did for the Defects in the Myocardial Insulin Signaling subheading.

•Current research: This section seem to be alright but for the heading 'Wilson Research Institute' there were no references cited.

•Glossary: Far too few terminologies were defined in this section.

Group7 : G protein (beta adrenergic)

•Introduction: A substantial amount of information is placed in this section giving readers a good overview of the topic. I can see that there were 7 linked references at the bottom. It will be better if they were placed right after the sentences that contained the corresponding information.

•History Pathway: Good and well referenced timeline.

•Gene Description: The dot point format of this section made it easier to understand. Having pictures would make it look more interesting and help the audience have a visual representation of what is being described.

•Receptor Structure: The last subheading for Beta-1 Adrenoceptor lacks a lot of information considering that this is the main receptor that your group has chosen to focus as mentioned in the introduction.

•Pathway and Normal Function: This is one of the well done parts of your project. Much research was done and all presented information were cited properly.

•Abnormal Function: This is the best section of your project. This section showed depth about the topic and related how the signaling pathway can cause the mentioned disorders. A suggestion to make this part more interesting would be gross images of the diseases.

•Glossary: This section is looking good. I suggest to bold the terms so it will be easier to read through this section.

This project is looking good. There are bits and pieces that has to be tied up. In addition, you need to put attention in the formatting of your page(especially for the abnormal function section). Nevertheless, it is looking good with a great deal of information about the topic. Lastly, you do not have a current research subheading.

Group8 : Leukocyte Extravasation

•Introduction: Introduction gives a extremely brief overview of the topic and no references were cited. This can be improved by going into a little bit more detail about the leukocyte extravasation.

•Pathway Section: This section was very clear but far too short with no references.

•History: This section has nothing yet.

•Normal Function: Shows understanding about the topic and was worded for audiences that are not familiar with the topic (which is a good point. that means you were able to relay the message in your own simple words). This section can be improved further by having references and checking the capitalization of some words. Also i think it will be better if the three numbered points under "brief explanation of the three steps of phagocytosis" under Leukocyte Activation were put in dot points. (just a suggestion)

•Abnormal Function: This section showed thorough research and is well referenced. As a suggestion, this section would look more engaging if there were photos.

•Proteins: Well researched section with proper references and appropriate images.

•New of Current Research: This section needs to be worked on. There is nothing under this section.

Group9 : p53

•Introduction: Good overview of the topic and information were well referenced.

•Pathway: Clear descriptions were given in this section however no reference were cited. There were three pathways mention which would look better if they were in different subheadings to give emphasis for each.

•Receptor: No information is under this section.

•Proteins: No information is under this section.

•History: Nicely placed on a table. Very good referencing (every period/date had a reference).

•Current research: This section only one current research. It would be good to have more than one current research about this topic considering that it is a 'hot' topic at the moment with a lot of research going on about this.

•Normal Function: Well researched section with good referencing. The function of p53 was well stated and having subheadings for it made the understanding for each easier. A good point about this section is that several proteins and cascade of reactions were mentioned but they were easy to understand and was not overwhelming as compared to how other groups presented this section. Good work.

•Abnormal Function: There was a reference cited for this section but was not properly referenced. This section should be worked on more and images will be good to add.


--Mark Hill 13:22, 17 May 2012 (EST) Your peer assessment shows that you have made some effort to work through each project with some specific comments for each project section. I am not sure about formatting of your comments and this suggests that you have cut and pasted from the formatted discussion page (where you added your group comments) or elsewhere (I will be comparing peer assessments). This made it difficult to read on this current page.

Lab 10

Stem Cells

Lab 11

Completion of the group project

Lab 12

1. Identify a current technique used in gene sequencing.

Next Generation Sequencing

2. Identify a recent cell biology research paper that has used microarray technology.

Myra O Villareal, Junkyu Han, Kenjiro Ikuta, Hiroko Isoda Mechanism of Mitf inhibition and morphological differentiation effects of hirsein A on B16 melanoma cells revealed by DNA microarray. J. Dermatol. Sci.: 2012, 67(1);26-36 PubMed 22564683

3. What aspect of the research findings were contributed by the microarray technique.

The paper used microarray technique to determine the response of B16 melanoma cells to Hirsein A(HA) and its effect in the down regulation of micropthalmia-associated transcription factor (MITF) gene. MITF transcription factors regulate the development of melanocytes. DNA microarray that was spotted with 265 genes for melanogenesis was assessed to determine the effect of HA in melanocytes.

The results showed that HA induced cells down regulates the MITF gene expression through intervention in the MAPK signaling pathway. This finding can be a helpful tool in the treatment of melanoma(skin cancer).