Archive for the ‘Induced Pluripotent Stem Cells’ Category

Pluripotent stem cells for improved reprogrammed Human …

Induced Pluripotent Stem Cells | Posted by admin
Feb 03 2019

Following the start of our recent collaboration with Phenocell, were pleased to be able to provide high quality Sebocytes developed from Human induced pluripotent stem cells (iPSC). Thanks to a perfectly standardized reprogramming protocol, they display lower batch to batch variability, allowing better reproducibility and accuracy of your experimental results.

Sebocytes have demonstrated their large potential to be unique tools for many life science research fields such as:

These cryopreserved reprogrammed pluripotent stem cells are available at low passage (P2), 2.106 cells/vial format and in 3 different phototypes (Caucasian, Asian and African). Developed from highly qualified Human iPS cells, each lot is validated, with a specific certificate of analysis, for all the following Sebocyte markers and specific functions.

Phenocells iPS-derived human Sebocytes display the typical epithelial morphology of primary sebocytes with heterogeneity in cell size due to lipid accumulation.

Phenocell Sebocytes from human induced pluripotent stem cells.

Expression of the two Sebocyte markers: MUC1 expressed in more than 80% of cells; KRT7 expressed in 80% of cells

Functional markers are strongly expressed after 5 days:

Evolution of specific markers in Sebocytes derived from Human induced pluripotent stem cells after 3 (d3) and 5 (d5) days in culture with the specific PhenoCULT-SEB culture medium, compared to primary keratinocytes (Ker).

KRT7 expression shows Sebocyte purity above 90%.

Black: isotype control; Red: anti-KRT7 antibody

Dose-dependent (up to 5-fold) lipid accumulation (Bodipy staining), response after a 24hr treatment with linoleic acid (LA)

PCi-SEB respond to a 96hr treatment with testosterone (10 M) by a 2-fold increase in lipid content. This response is significantly inhibited by the 5-reductase inhibitor Finasteride (10 M).

Interested in these Sebocytes developed from Human induced pluripotent stem cells ?

Get in touch by leaving a comment below, Ill be pleased to get back to you to discuss your projects and needs.

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Pluripotent stem cells for improved reprogrammed Human ...

Induced Pluripotent Stem Cell Market Is Expected to Reach US …

Induced Pluripotent Stem Cells | Posted by admin
Feb 03 2019

New York, NY -- (SBWIRE) -- 02/02/2019 -- The healthcare industry has been focusing on excessive research and development in the last couple of decades to ensure that the need to address issues related to the availability of drugs and treatments for certain chronic diseases is effectively met. Healthcare researchers and scientists at the Li Ka Shing Faculty of Medicine of the Hong Kong University have successfully demonstrated the utilization of human induced pluripotent stem cells or hiPSCs from the skin cells of the patient for testing therapeutic drugs.

To know key findings Request Sample Report @: https://www.persistencemarketresearch.com/samples/17968

The success of this research suggests that scientists have crossed one more hurdle towards using stem cells in precision medicine for the treatment of patients suffering from sporadic hereditary diseases. iPSCs are the new generation approach towards the prevention and treatment of diseases that takes into account patients on an individual basis considering their genetic makeup, lifestyle, and environment. Along with the capacity to transform into different body cell types and same genetic composition of the donors, hiPSCs have surfaced as a promising cell source to screen and test drugs.

In the present research, hiPSC was synthesized from patients suffering from a rare form of hereditary cardiomyopathy owing to the mutations in Lamin A/C related cardiomyopathy in their distinct families. The affected individuals suffer from sudden death, stroke, and heart failure at a very young age. As on date, there is no exact treatment available for this condition. This team in Hong Kong tested a drug named PTC124 to suppress specific genetic mutations in other genetic diseases into the iPSC transformed heart muscle cells. While this technology is being considered as a breakthrough in clinical stem cell research, the team at Hong Kong University is collaborating with drug companies regarding its clinical application.

The unique properties of iPS cells provides extensive potential to several biopharmaceutical applications. iPSCs are also used in toxicology testing, high throughput, disease modeling, and target identification. This type of stem cell has the potential to transform drug discovery by offering physiologically relevant cells for tool discovery, compound identification, and target validation. A new report by Persistence Market Research (PMR) states that the global induced pluripotent stem or iPS cell market is expected to witness a strong CAGR of 7.0% from 2018 to 2026. In 2017, the market was worth US$ 1,254.0 Mn and is expected to reach US$ 2,299.5 Mn by the end of the forecast period in 2026.

Request for Report Methodology @: https://www.persistencemarketresearch.com/methodology/17968

Customization to be the Key Focus of Market Players

Due to the evolving needs of the research community, the demand for specialized cell lines have increased to a certain point where most vendors offering these products cannot depend solely on sales from catalog products. The quality of the products and lead time can determine the choices while requesting custom solutions at the same time. Companies usually focus on establishing a strong distribution network for enabling products to reach customers from the manufacturing units in a short time period.

Get full Report Now: https://www.persistencemarketresearch.com/checkout/17968

Entry of Multiple Small Players to be Witnessed in the Coming Years

Several leading players have their presence in the global market; however, many specialized products and services are provided by small and regional vendors. By targeting their marketing strategies towards research institutes and small biotechnology companies, these new players have swiftly established their presence in the market.

About Persistence Market Research Persistence Market Research (PMR) is a third-platform research firm. Our research model is a unique collaboration of data analytics and market research methodology to help businesses achieve optimal performance. To support companies in overcoming complex business challenges, we follow a multi-disciplinary approach. At PMR, we unite various data streams from multi-dimensional sources. By deploying real-time data collection, big data, and customer experience analytics, we deliver business intelligence for organizations of all sizes.

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Induced Pluripotent Stem Cell Market Is Expected to Reach US ...

Pluripotent Stem Cell Flow Kit (FMC001): R&D Systems

Induced Pluripotent Stem Cells | Posted by admin
Feb 03 2019

H/M Pluripotent Stem Cell Multi-Color Flow Cytometry Kit Summary Kit Summary For the verification of stem cell pluripotency using four established markers. Key Benefits

Identifying the sources of experimental variability is an important consideration in stem cell research where experiments are costly and time-consuming. A potential source of significant variability arises from the starting population of stem cells which can undergo phenotypic changes in culture. SeeDetails

Changes in stem cell potency over time can give rise to large inter-assay errors and/or contradictory data. The Human/Mouse Pluripotent Stem Cell Multi-Color Flow Cytometry Kit offers users an efficient and quantitative method to verify the pluripotency of cells by flow cytometry. Data obtained using this kit can identify and minimize experimental errors introduced by variations in the starting population of cells.

The Human/Mouse Pluripotent Stem Cell Multi-Color Flow Cytometry Kit includes four fluorochrome-conjugated primary antibodies, isotype controls and buffers to fix, permeabilize, and wash cells. SeeDetails

Store at 2 C to 8 C in the dark. Use within 6 months of receipt.

Verification of Human BG01V Embryonic Stem Cell Pluripotency by Multi-Color Flow Cytometry. BG01V human embryonic stem cells were stained using reagents included in the Human/Mouse Pluripotent Stem Cell Multi-Color Flow Cytometry Kit (Catalog# FMC001). Cells were simultaneously analyzed for expression of pluripotent markers including SSEA-1, SSEA-4, Oct-3/4, and SOX2 by flow cytometry. A. Flow cytometry data shows that 91.9% of BG01V human embryonic stem cells are positive for both Oct-3/4 and SSEA4 expression. B. Flow cytometry data shows that 88.5% of BG01V human embryonic stem cells are positive for SSEA-4 and negative for SSEA-1, a phenotype consistent with human embryonic stem cells. C. Flow cytometric analysis shows that BG01V human embryonic stem cells express the pluripotent marker SOX2.

Verification of Mouse D3 Embryonic Stem Cell Pluripotency by Multi-Color Flow Cytometry. Mouse D3 embryonic stem cells were stained using reagents included in the Human/Mouse Embryonic Stem Cell Multi-Color Flow Cytometry Kit (Catalog#FMC001). Cells were analyzed for expression of pluripotent markers including SSEA-1, SSEA-4, Oct-3/4, and SOX2 by flow cytometry. A. Flow cytometric analysis shows that 91.1% of mouse D3 embryonic stem cells are positive for both Oct-3/4 and SSEA1 expression. B. Flow cytometric analysis data shows that 82.6% of mouse D3 embryonic stem cells are positive for SSEA-1 and negative for SSEA-4 a phenotype consistent with mouse embryonic stem cells. C. Flow cytometric analysis shows that mouse D3 embryonic stem cells express the pluripotent marker SOX2.

BG01V human embryonic stem cells are licensed from ViaCyte, Inc.

Stability & Storage

Store the unopened product at 2 - 8 C. Do not use past expiration date.

Embryonic stem (ES) cells are pluripotent stem cells derived from the inner cell mass of pre-implantation embryos. Induced pluripotent stem (iPS) cells can be generated by somatic cell reprogramming following the exogenous expression of specific transcription factors (Oct-3/4, KLF4, SOX2, and c-Myc). These cell types are capable of unlimited, undifferentiated proliferation in vitro and still maintain the capacity to differentiate into a wide variety of somatic cells. In this capacity, pluripotent stem cells have widespread clinical potential for the treatments of heart disease, diabetes, spinal cord injury, and a variety of neurodegenerative disorders.

R&D Systems offers a wide range of products to support pluripotent stem cell culture and differentiation. Mouse embryonic fibroblasts may be used to maintain and expand pluripotent stem cells in an undifferentiated state. We also offer defined culture media, which are specifically optimized for use with human or rodent pluripotent stem cells. In addition, R&D Systems offers a variety of products to assess differentiation status and identify specific stem cell types of interest, including panels of marker antibodies, primer pairs, multi-color flow cytometry kits, and specialized verification kits.

Alternate Names

Pluripotent Stem Cells

WARNING: This product can expose you to chemicals including formaldehyde and methanol, which are known to the State of California to cause cancer and reproductive toxicity with developmental effects. For more information, go to http://www.P65Warnings.ca.gov.

Refer to the product datasheet for complete product details.

Reagents Supplied in the Human/Mouse Embryonic Stem Cell Multi-Color Flow Cytometry Kit (Catalog # FMC001)

Intracellular Staining Protocol with Simultaneous Fixation/Permeabilization

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

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Pluripotent Stem Cell Flow Kit (FMC001): R&D Systems

induced pluripotent stem cell | The Science of Parkinson’s

Induced Pluripotent Stem Cells | Posted by admin
Feb 03 2019

Parkinsons is a neurodegenerative condition. This means that cells in the brain are being lost over time. Any cure for Parkinsons is going to require some form of cell replacement therapy introducing new cells that can replace those that were lost.

Cell transplantation represents one approach to cell replacement therapy, and this week we learned that the Japanese regulatory authorities have given the green light for a new cell transplantation clinical trial to take place in Kyoto.

This new trial will involve cells derived from induced pluripotent stem cells (or IPS cells).

Intodays post we will discuss whatinduced pluripotent stem cells are, what previous research has been conducted on these cells, and what we know about the new trial.

Source:Glastone Institute

The man in the image above is ProfShinya Yamanaka.

Hes a rockstar in the biomedical research community.

ProfYamanaka is the director ofCenter forinduced Pluripotent Stem CellResearch and Application(CiRA); and a professor at theInstitute for Frontier Medical SciencesatKyoto University.

But more importantly, in 2006 he published a research report that would quite literally change everything.

In that report, he demonstrated a method by which someonecould take a simple skin cell (called a fibroblast), grow it in cell culture for a while, and then re-programit so that it would transform into a stem cell a cell that is capable of becoming any kind of cell in the body.

The transformed cells were calledinduced pluripotent stem (IPS) cell pluripotent meaning capable of any fate.

It was an amazing feat that made the hypothetical idea of personalised medicine suddenly very possible take skin cells from anyone with a particular medical condition, turn them into whatever cell type you like, and then either test drugs on those cells or transplant them back into their body (replacing the cells that have been lost due to the medical condition).

Personalised medicine with IPS cells. Source:Bodyhacks

IPS cells are now being used all over the world, for all kinds of biomedical research. And many research groups are rushing to bring IPS cell-based therapies to the clinic in the hope of providing the long sort-after dream of personalised medicine.

This week the Parkinsons community received word that the Pharmaceuticals and Medical Devices Agency (PMDA) the Japanese regulatory agency that oversees clinical trials have agreed for researchers at Kyoto University to conduct a cell transplantation trial for Parkinsons, using dopamine neurons derived from IPS cells. And the researchers are planning to begin their study in the next month.

In todays post we are going to discuss this exciting development, but we should probably start at the beginning with the obvious question:

What exactly is an IPS cell?

Continue reading

New research provides some interesting insight into particular cellular functions and possibly sleep issues associated with Parkinsons.

Researchers in Belgium have recently published interesting findings that a genetic model of Parkinsons exhibits sleep issues, which are not caused by neurodegeneration, but rather neuronal dysfunction. And as a result, they were able to treat it in flies at least.

In todays post, we will review this new research and consider its implications.

Source:Dlanham

I am a night owl.

One that is extremely reluctant to give up each day to sleep. There is always something else that can be done before going to bed. And I can often be found pottering around at 1 or 2am on a week night.

As a result of this foolish attitude, I am probably one of the many who live in a state of sleep deprivation.

I am a little bit nervous about doing the spoon test:

But I do understand that sleep is very important for our general level of health and well being. And as a researcher on the topic, I know that sleep complications can be a problem for people with Parkinsons.

What sleep issues are there for people with Parkinsons?

Continue reading

This week a group of scientists have published an article which indicates differences between mice and human beings, calling into question the use of these mice in Parkinsons disease research.

The results could explain way mice do not get Parkinsons disease, and theymay also partly explain why humans do.

In todays post we will outline the new research, discuss the results, and look at whether Levodopa treatment may (or may not) be a problem.

The humble lab mouse. Source: PBS

Much of our understanding of modern biology is derived from the lower organisms.

From yeast to snails (there is a post coming shortly on a snail model of Parkinsons disease I kid you not) and from flies to mice, a great deal of what we know about basic biology comes from experimentation on these creatures. So much in fact that many of our current ideas about neurodegenerative diseases result from modelling those conditions in these creatures.

Now say what you like about the ethics and morality of this approach, these organisms have been useful until now. And I say until now because an interesting research report was released this week which may call into question much of the knowledge we have from the modelling of Parkinsons disease is these creatures.

You see, heres the thing: Flies dont naturally develop Parkinsons disease.

Nor do mice. Or snails.

Or yeast for that matter.

So we are forcing a very un-natural state upon the biology of these creatures and then studying the response/effect. Which could be giving us strange results that dont necessarily apply to human beings. And this may explain our long history of failed clinical trials.

We work with the best tools we have, but it those tools are flawed

What did the new research report find?

This is the study:

Title: Dopamine oxidation mediates mitochondrial and lysosomal dysfunction in Parkinsons disease Authors: Burbulla LF, Song P, Mazzulli JR, Zampese E, Wong YC, Jeon S, Santos DP, Blanz J, Obermaier CD, Strojny C, Savas JN, Kiskinis E, Zhuang X, Krger R, Surmeier DJ, Krainc D Journal: Science, 07 Sept 2017 Early online publication PMID:28882997

The researchers who conducted this study began by growing dopamine neurons a type of cell badly affected by Parkinsons disease from induced pluripotent stem (IPS) cells.

What are induced pluripotent stem cells?

Continue reading

Two months ago a research report was published in the scientific journal Nature and it caused a bit of a fuss in the embryonic stem cell world.

Embryonic stem (ES) cells are currently being pushed towards the clinic as a possible source of cells for regenerative medicine. But this new report suggested that quite a few of the embryonic stem cells being tested may be carrying genetic variations that could be bad. Bad as in cancer bad.

In this post, I will review the study and discuss what it means for cell transplantation therapy for Parkinsons disease.

Source: Medicalexpress

For folks in the stem cell field, the absolute go-to source for all things stem cell related isProf Paul Knoepflers blog The Niche. From the latest scientific research to exciting new stem cell biotech ventures (and even all of the regulatory changes being proposed in congress), Pauls blog is a daily must read for anyone serious about stem cell research. He has his finger on the pulse and takes the whole field very, very seriously.

Prof Paul Knoepfler during his TED talk.Source: ipscell

For a long time now, Paul has been on a personal crusade. Like many others in the field (including yours truly), he has been expressing concern about the unsavoury practices of the growing direct-to-consumer, stem cell clinic industry. You may have seen him mentioned in the media regarding this topic (such as this article).

The real concern is that while much of the field is still experimental, many stem cell clinics are making grossly unsubstantiated claims to draw in customers. From exaggerated levels of successful outcomes (100% satisfaction rate?) all the way through to talking about clinical trials that simply do not exist.The industry is badly (read: barely) regulated which is ultimately putting patients at risk (one example: three patients were left blind after undergoing an unproven stem cell treatment click here to read more on this).

While the stem cell research field fully understands and appreciates the desperate desire of the communities affected by various degenerative conditions, there has to be regulations and strict control standards that all practitioners must abide by. And first amongst any proposed standards should be that the therapy has been proven to be effective for a particular condition in independently audited double blind, placebo controlled trials. Until such proof is provided, the sellers of such products are simply preying on the desperation of the people seeking these types of procedures.

Continue reading

Last weekscientists in Sweden published researchdemonstrating a method by which the supportive cells of the brain (called astrocytes) can be re-programmed into dopamine neurons in the brain of a live animal!

It was a reallyimpressive trick and it could have major implications for Parkinsons disease.

In todays post is a long read, but in it we will review the research leading up to the study, explain the science behindthe impressive feat, and discuss where things go from here.

Different types of cells in the body. Source: Dreamstime

In your body at this present moment in time, there is approximately 40 trillion cells (Source).

The vast majority of those cells have developedinto mature types of cell and they are undertaking veryspecific functions. Muscle cells, heart cells, brain cells all working together in order to keep you verticaland ticking.

Now, once upon a time we believed that the maturation (or the more technical term: differentiation) of a cell was a one-way street. That is to say, once acellbecame what it was destined to become, there was no going back. This was biological dogma.

Then aguy in Japan did something rather amazing.

Who is he and what did he do?

This is ProfShinya Yamanaka:

ProfShinya Yamanaka. Source: Glastone Institute

Hes a rockstar in the scientific research community.

ProfYamanaka is the director of Center for induced Pluripotent Stem Cell Research and Application(CiRA); and a professor at theInstitute for Frontier Medical Sciences at Kyoto University.

But more importantly, in 2006 he published a research reportdemonstrating how someonecould take a skin cell and re-programit so that was now a stem cell capable of becoming any kind of cell in the body.

Heres the study:

Title: Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Authors: Takahashi K, Yamanaka S. Journal: Cell. 2006 Aug 25;126(4):663-76. PMID: 16904174 (This article is OPEN ACCESS if you would like to read it)

Shinya Yamanakas team started with the hypothesis that genes which are important to the maintenance of embryonic stem cells (the cells that give rise to all cells in the body) might also be able to cause an embryonic state in mature adult cells. They selected twenty-four genes that had been previously identified as important in embryonic stem cells to test this idea. They used re-engineered retroviruses to deliver these genes to mouse skin cells. The retroviruses were emptied of all their disease causing properties, and could thus function as very efficient biological delivery systems.

The skin cells were engineered so that only cells in which reactivation of the embryonic stem cells-associated gene, Fbx15, would survive the testing process. If Fbx15 was not turned on in the cells, they would die. When the researchers infected the cells with all twenty-four embryonic stem cells genes, remarkably some of the cells survived and began to divide like stem cells.

In order to identify the genes necessary for the reprogramming, the researchers began removing one gene at a time from the pool of twenty-four. Through this process, they were able to narrow down the most effective genes to justfour: Oct4, Sox2, cMyc, and Klf4, which became known as the Yamanaka factors.

This new type of cell is called an induced pluripotent stem (IPS) cell pluripotent meaning capable of any fate.

The discovery of IPS cells turned biological dogma on its head.

And in acknowledgement of this amazing bit of research, in 2012 ProfYamanaka and Prof John Gurdon (University of Cambridge)were awarded the Nobel prize for Physiology and Medicinefor the discovery that mature cells can be converted back to stem cells.

Prof Yamanaka and Prof Gurdon. Source: UCSF

Prof Gurdon achieved the feat in 1962 when he removed the nucleus of a fertilised frog egg cell and replaced it with the nucleus of a cell taken from a tadpoles intestine. The modified egg cell then grew into an adult frog! This fascinatingresearchproved that the mature cell still contained the genetic information needed to form all types of cells.

EDITORS NOTE: We do not want to be accused of taking anything away from Prof Gurdons contribution to this field (which was great!) by not mentioning his efforts here. For the sake of saving time and space, we are focusing onProf Yamanakas research as it is more directly related to todays post.

Making IPS cells. Source: learn.genetics

Link:
induced pluripotent stem cell | The Science of Parkinson's

What Are Induced Pluripotent Stem Cells? | Intro to the …

Induced Pluripotent Stem Cells | Posted by admin
Jan 26 2019

The team of Japanese doctors, led by cardiac surgeon Yoshiki Sawa at Osaka University, will useiPS cells to create a sheet of 100 million heart-muscle cells.It will be the second clinical application of iPS cells in Japan and third worldwide (RIKEN, Cynata Therapeutics, and now, Osaka University).

The growing popularity of iPSC technology has also been attracting investments from the commercial sector. Notably, in December 2016, Bayer AG and Versant Ventures formed a start-up named BlueRock Therapeutics focused on iPSCs therapy. The company raised funding of USD $225 million, the largest iPSC financing round ever.

The largest company manufacturing iPS cells isCellular Dynamics International, a Fujifilm company. The company is widely known as Fujifilm CDI.

Fujifilm CDI manufactures biologically relevant human cells derived from iPS cells. Its iCell and donor-specific MyCell Products are highly pure, highly reproducible, and available in industrial quantity to enable drug discovery, toxicity testing, stem cell banking, and cell therapy development.

Within Europe, the largest iPS cell developer and manufacturer is Ncardia, a company formed in September 2017 by the merger of Axiogenesis and Pluriomics.Ncardia is the largest supplier in Europe and the second largest iPS cell company in the world after Fujifilm CDI.

Ncardia is a private company with operations in Europe and the US that produces and commercializes high-quality, fully functional hiPSC derived cardiovascular and neuronal cell types. It also develops electrophysiology, biochemistry,and contraction based assays to support drug development and discovery.

There are also dozens of other suppliers of iPS cell lines, differentiated cell types, kits, assays, reprogramming services, and more.

Today, methods for commercializing iPS cells are still being explored, as clinical studies investigating them remain low in number. One of the greatest challenges is to establish standards across the industry for cell quality and functionality in order to protect patient safety.

To learn more about iPS cells, view the video below:

If you found this blog valuable, subscribe to BioInformants stem cell industry updates.

As the first and only market research firm to specialize in the stem cell industry, BioInformant research is cited by The Wall Street Journal, Xconomy, AABB, and Vogue Magazine. Bringing you breaking news on an ongoing basis, we encourage you to join more than half a million loyal readers, including physicians, scientists, executives, and investors.

Do you think iPS cells are safe for use within cell therapy? What do you see as their pros and cons? Leave your thought in the comments section below.

Up Next:Worlds First Clinical Application of iPS Cells for Cardiac Disease

Editors Note: This post was originally published on June 25, 2018, and has been updated for quality and relevancy.

Read more from the original source:
What Are Induced Pluripotent Stem Cells? | Intro to the ...

Pluripotent Stem Cells – Stemcell Technologies

Induced Pluripotent Stem Cells | Posted by admin
Dec 30 2018

Pluripotent Stem Cells

Few areas of biology currently garner more attention than the study of human pluripotent stem cells (hPSCs). This interest has arisen because of their potential to form the basis of cellular therapies for diseases affecting organ systems with limited regenerative capacity, to provide enhanced systems for drug screening and toxicity testing as well as to gain insight into early human development. There are currently two major methods for generating cells with pluripotent properties. The first involves isolating the inner cell mass from an early human blastocyst and culturing the resulting cells in appropriate culture conditions (see below) to generate human embryonic stem cells (hESCs).1 The second involves artificially expressing key developmental transcription factors in somatic cell types, which, with the appropriate culture conditions, causes the cells to be reprogrammed into induced pluripotent stem cells (iPSCs).24

Much effort has been dedicated to understanding the transcriptional state of undifferentiated pluripotent stem cells. For example, OCT-3/4,5 KLF-4, SOX2,6 and NANOG7,8 have been shown to be central to the specification of pluripotent stem cell identity due to their unique expression patterns and their essential roles in early development. These efforts, along with others, enabled the discovery of defined factors for the reprogramming of somatic cells. The specific molecular processes by which somatic cells are reprogrammed into pluripotent stem cells are not understood, although recent findings suggest it is a stepwise process,9,10 and that the stochastic nature of reprogramming can be explained in part by the multiple molecular and genetic events required for full reprogramming to occur.11 Current research focuses on improving reprogramming efficiency by better understanding several variables in the process of reprogramming: (1) the choice of factors used; (2) the delivery methods; (3) the target cell type; (4) the timing and levels of factor expression; and (5) the culture conditions. In addition to this, methods to identify and characterize truly reprogrammed pluripotent cells are also important.12

The original cocktail of factors described by Yamanaka13, OCT4, SOX2, c-MYC and KLF-4, continue to be the major factors that are used for reprogramming. Originally, delivery of the factors was achieved through the use of viruses that integrated into the genome. However, concerns over the clinical use of these cells and the potential for insertional mutagenesis have led to the exploration of non-integrating methods of factor delivery including transient transfection,14 non-integrating viral approaches15 and protein transduction.16 Other recent methods such as the use of polycystronic minicircles,17 synthetic mRNA,18 self-replicating RNA,19 RNA based viruses such as Sendai,20 and synthetic microRNA21 have also been shown to be successful. Particularly exciting are recent research efforts to identify small molecules that replace some of these factors by either modifying genome methylation patterns or inhibiting key signaling pathways.22,23 The ultimate goal of this research is to define stepwise protocols by which cells can be fully reprogrammed solely by chemical means.

The choice of cell type to use for reprogramming is based on accessibility of tissue samples, genetic make-up of the target cells, and reprogramming efficiency. Skin-derived dermal fibroblast cells and peripheral blood cells are the most commonly used cell types due to the limited invasiveness of sample collection and availability of banked tissue samples representing a variety of diseases. Peripheral blood cell types have varying and often reciprocal efficiencies of reprogramming versus frequency in blood. For example, CD34+ hematopoietic stem and progenitors have relatively high reprogramming efficiencies24 but are rare in circulating blood (0.01-0.1%).25 In contrast, T- and B-cells are more frequent and have acceptable reprogramming efficiency,26 but are less ideal as target cells for reprogramming due to TCR and IgG gene rearrangements that may affect the downstream function of hiPSCs generated from them.27 Thus, peripheral blood represents a promising and readily available source of cells for reprogramming.

Interestingly, not all cell types require all four factors to be delivered in order to successfully reprogram cells. For example, as neural stem cells endogenously express SOX2, KLF-4, and c-MYC, they were able to be reprogrammed solely through transduction of OCT-4.28 Cell types also appear to have different reprogramming efficiencies. In mice, stomach and liver cells appear to be reprogrammed more efficiently and completely than fibroblasts.29 Similarly, the reprogramming of human adipocytes is ~20-fold more efficient than fibroblasts and have the added advantage of being a readily available source of cells.30,31 As a final consideration when choosing a starting cell type, several reports have noted retained gene expression from the parental cell types32 and have also shown that the epigenetic state is predictive of the original somatic cell type.33 Such epigenetic memory may increase the propensity of iPS cell lines to differentiate to the original cell lineage/type.33-35

A common phenomenon observed during reprogramming is the emergence of partially reprogrammed colonies which are usually associated with continued expression of the reprogramming factors. These cells exhibit a range of phenotypes but often fail tests of fully pluripotent cells.37 Chan et al.38 showed that while overall reprogramming efficiency was lower in feeder-free conditions, the only types of cells that emerged were fully reprogrammed cells. This indicates the importance of culture conditions in the process of reprogramming. We developed feeder-free, defined and xenofree media for reprogramming fibroblasts (TeSR-E7) or blood cells (ReproTeSR), which provide recognizable hiPSC colonies with less differentiated or partially reprogrammed background cell growth.

Initial methods to culture hESCs were modeled on techniques originally developed to culture mouse ESCs (mESCs).39,40 These techniques involved culture on a layer of mitotically inactivated mouse embryonic fibroblasts (MEFs, or feeder cells) in medium supplemented with 20% fetal bovine serum (FBS). In these conditions, hESC lines could be propagated indefinitely with retention of their pluripotent properties.1 From the initial development of these culture conditions, it was realized that the continued use of feeders and animal-derived components in hESC cultures would hinder the development of clinical applications due to: a) the presence of immunogenic material; b) the risk of transmitting animal virus or prion material; and c) difficulty with quality control of these undefined components.

Subsequently, improvements to these procedures have largely focused on removing the undefined and non-human components. Several groups have developed culture conditions for hESCs that are, to various degrees, serum- and MEF-free. It was discovered that, in MEF-dependent conditions, serum could be replaced with Knock-Out Serum Replacement,41 a commercially available serum substitute. Xu et al. reported a culture system that utilized Matrigel as a culture matrix and MEF-conditioned medium (consisting of serum replacement and basic fibroblast growth factor, bFGF) that allowed hESCs to be cultured without direct contact with feeders.42 Another approach to MEF removal from the culture system was to replace them with human feeders.43 As the feeders are of human origin, the possibility of the transmission of foreign pathogens is limited, but unfortunately the secreted factors are still undefined and subject to large variation between batches.

True feeder-free culture has been achieved using an extracellular matrix surface coating on the cultureware, and a combination of transforming growth factor- (TGF-) and bFGF or high levels of bFGF alone44,45 together with a serum replacement in the medium. A number of publications have described defined xeno- or feeder-free media formulations for the maintenance of hESCs.46-49

The TeSR family of defined and serum-free media for feeder-free culture of hPSCs includes mTeSR1, TeSR2, and TeSR-E8. mTeSR1 was developed by Dr. Tenneille Ludwig and colleagues at the WiCell Research Institute (Madison, WI) and supports longterm, feeder-free culture of hESCs and hiPSCs.49 The formulation of mTeSR1 includes key factors that support pluripotency including bFGF, TGF-, -aminobutyric acid (GABA), pipecolic acid and lithium chloride, as well as bovine serum albumin (BSA). mTeSR1 is now the most widely-published feeder-free medium, used in over 800 peer-reviewed publications. TeSR2 is a more defined medium, based on the xeno-free formulation from the same group, containing recombinant HSA.47 The WiCell Research Institute also developed a low protein, highly defined culture medium for hPSCs. This medium, TeSR-E8, contains only the most essential components required for maintenance providing a simpler medium for the culture of pluripotent stem cells.

hPSCs differ at the molecular and functional level from mESCs and are considered to more closely resemble post-implantation mouse epiblast stem cells (EpiSCs). mESCs and conventional hPSCs exhibit distinct gene expression patterns and different requirements in culture.50,51 Specifically, mESCs are maintained by inhibiting MEK/ERK signaling, activating WNT signaling (by GSK3 inhibition), and stimulating with the leukemia inhibitory factor (LIF) cytokine, while hPSCs or mouse EpiSCs are cultured in FGF and Activin and are not responsive to LIF.50 Several recent studies have identified conditions capable of maintaining hPSCs in a ground state resembling mESCs as opposed to the primed state that hPSCs are traditionally maintained in.51-53

A lot of effort has focused on finding surface matrices that are more defined than Matrigel . Two of the more promising approaches are synthetic peptides chemically linked to the cultureware,54,55 and recombinant proteins that interact with specific integrins and cell adhesion molecules.56,57 A new defined surface, Vitronectin XF, was developed and manufactured by Primorigen Biosciences and has been commercially released by STEMCELL Technologies. Vitronectin XF can be used with mTeSR1 or, TeSR2 or TeSR-E8 for a xeno-free culture system.

Because of their differentiation potential, it is hoped that hPSCs may form the basis of cellular therapies where tissue damage or malfunction is severe and irreversible. Cardiovascular diseases, type-1 diabetes, spinal cord injury, and Parkinsons disease are examples of diseases where it is hoped that hPSC-based therapies will provide a cure. Techniques have been developed to differentiate hPSCs into a variety of adult cell types including hematopoietic,58-60 cardiac,61,62 neural,63-65 pancreatic,66-71 retinal pigmented epithelia72,73 and osteogenic lineages.74 However, a number of obstacles currently impede the clinical application of hPSC-based therapies. At present, only limited testing of hPSC-derived cells has been performed to ensure full maturation and functionality of differentiated cells. Furthermore, protocols for the differentiation of hPSCs to functionally relevant progeny are generally inefficient, resulting in low differentiated cell yields and contamination by other cell types as a result of aberrant differentiation. Of greater concern is the possibility of the persistence of undifferentiated hPSCs in transplanted populations which may result in teratomas.75,76

Further concerns surround the possibility of immune rejection of transplanted cells either due to the expression of different major histocompatibility complex antigens on donor cells77 or from the expression of foreign antigens as a result of culturing in animal products.78 Using patient-specific iPSCs for cellular therapies would circumvent the need for histocompatibility matching. And while the potential of rejection due to foreign animal antigens remains controversial,78,79 much effort is being devoted to developing xeno-free culture media and matrices for hPSC expansion and differentiation. STEMCELL Technologies offers the STEMdiff suite of defined and feeder-free products for efficient differentiation of hPSCs to cells of all three lineages.

Clinical application of hPSC-based therapies are moving closer towards becoming a reality. For example, the group led by Dr. Masayo Takahashi at RIKEN, Japan, recently started treating the first patients with age-related macular degeneration using autologous hiPSC-derived retinal pigmented epithelial cells. Similarly, early stage clinical trials are ongoing in the U.S. andthe E.U. by Advanced Cell Technologies to utilize hESC-derived retinal pigmented epithelial cells to treat Stargarts macular dystrophy, a degenerative eye disease that causes blindness in children. Finally, ViaCyte has recently received FDA acceptance of IND for the candidate hESC-derived beta cell replacement therapy for type 1 diabetes and will begin phase 1 clinical trials soon.

The most immediate impacts are likely to be gained from the use of hPSCs in the fields of drug development or toxicity testing. It has been estimated that the cost of bringing a new drug to market through development, clinical trials and FDA approval can be upwards of 800 million to 1.3 billion USD.80 Furthermore the number of drugs that are ultimately successful is very low, and many drugs fail at the phase II or III clinical trial stages due to unexpected toxicities, after large investments have been put into their development. Given these costs and the high risk assumed by pharmaceutical companies, there are great advantages to having access to large numbers of biologically relevant human cells for early testing and screening. hPSCs in their undifferentiated state may be useful to identify teratogenic or toxic effects of potential compounds. Incorporating compounds into defined differentiation protocols may identify candidates that potentiate or skew differentiation towards a beneficial outcome. The potential to generate large numbers of end stage cells such as neurons and cardiomyocytes will ultimately provide directly relevant cell types for drugs being developed for cardiovascular or neurodegenerative disorders. In addition, the generation of cardiomyocytes and hepatocytes may be directly relevant to toxicity measurements. Finally, disease-specific iPSCs made by reprogramming relevant cell types from patients has the potential of revealing not only fundamental biological defects but also providing potentially unlimited cells with which to investigate potential therapeutic approaches.

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Pluripotent Stem Cells - Stemcell Technologies

ATCC-HYR0103 Human Induced Pluripotent Stem (IPS) Cells ATCC …

Induced Pluripotent Stem Cells | Posted by admin
Dec 30 2018

Complete Growth Medium

ATCC iPSCs have been adapted to feeder- and serum-free culture conditions.

The base medium for this cell line is Pluripotent Stem Cell SFM XF/FF (ATCC No. ACS-3002) which is a ready-to-use medium for serum-free and feeder-free iPSC culture.

Cell culture dishes are coated with CellMatrix Basement Membrane Gel (ATCC No. ACS-3035) to provide a surface for the attachment of iPSCs.

Coating Procedure:

Dilute CellMatrix in DMEM:F12 to a working concentration of 150 g/mL. For instance, if the protein concentration of CellMatrix (on certificate of analysis) is 14 mg/mL, then: (4 mL) x (0.15 mg/mL)/(14 mg/mL) = 0.043 mL. Therefore, add 43 L CellMatrix directly in 4 mL cold DMEM: F-12 Medium

ROCK Inhibitor Y27632 is not necessary each time the culture medium is changed. It is required when cells are recovering from thaw on CellMatrix Gel-coated dishes containing 5 mL Pluripotent Stem Cell XF/FF medium/6-cm dish.

This protocol is designed to passage stem cell colonies cultured in a 6 cm dish, using Stem Cell Dissociation Reagent (ATCC ACS-3010) to detach the cell colonies. The recommended spilt ratio is 1:4. Volumes should be adjusted according to the size and number of the tissue culture vessels to be processed.

Lyophilized proteins tend to be hygroscopic. Bring the vial of Stem Cell Dissociation Reagent to room temperature before opening. The vial should not be cool to the touch. Once opened, the lyophilized material should be stored desiccated. The specific activity of the reagent is found on the certificate of analysis. Dissolve the appropriate amount of Stem Cell Dissociation Reagent in DMEM: F-12 Medium to prepare a 0.5 U/mL working solution.

Note: Addition of ROCK inhibitor has been shown to increase the survival rate. The use of ROCK inhibitor may cause a transient spindle-like morphology effect on the cells. However, the colony morphology will recover after subsequent media change without ROCK inhibitor.

For optimal results, cryopreserve stem cell colonies when the cell cultures are 80% confluent. This protocol is designed to cryopreserve stem cell colonies cultured in a 6 cm dish.

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Induced Pluripotent Stem Cells Market Report Research …

Induced Pluripotent Stem Cells | Posted by admin
Dec 30 2018

Pluripotent stem cells are embryonic stem cells that have the potential to form all adult cell types and help in repairing of damaged tissues in the human body. An induced pluripotent stem cells, or iPSCs, are taken from any tissue (usually skin or blood) from a child or an adult and is genetically modified to behave like pluripotent stem cells or embryonic stem cells. iPSCs market is in emerging state mainly due to its ability to make any cell or tissue the body might need to encounter wide range of diseases like diabetes, spinal cord injury, leukaemia or heart disease, these cells can potentially be customized to provide a perfect genetic match for any patient. In 2018, the global Induced Pluripotent Stem Cells market size was xx million US$ and it is expected to reach xx million US$ by the end of 2025, with a CAGR of xx% during 2019-2025.

This report focuses on the global Induced Pluripotent Stem Cells status, future forecast, growth opportunity, key market and key players. The study objectives are to present the Induced Pluripotent Stem Cells development in United States, Europe and China.

The key players covered in this study Fujifilm Holding Corporation Astellas Pharma Fate Therapeutics Bristol-Myers Squibb Company ViaCyte Celgene Corporation Aastrom Biosciences Acelity Holdings StemCells Japan Tissue Engineering Organogenesis

Market segment by Type, the product can be split into Hepatocytes Fibroblasts Keratinocytes Amniotic Cells Others

Market segment by Application, split into Academic Research Drug Development And Discovery Toxicity Screening Regenerative Medicine

Market segment by Regions/Countries, this report covers United States Europe China Japan Southeast Asia India Central & South America

The study objectives of this report are: To analyze global Induced Pluripotent Stem Cells status, future forecast, growth opportunity, key market and key players. To present the Induced Pluripotent Stem Cells development in United States, Europe and China. To strategically profile the key players and comprehensively analyze their development plan and strategies. To define, describe and forecast the market by product type, market and key regions.

In this study, the years considered to estimate the market size of Induced Pluripotent Stem Cells are as follows: History Year: 2014-2018 Base Year: 2018 Estimated Year: 2019 Forecast Year 2019 to 2025 For the data information by region, company, type and application, 2018 is considered as the base year. Whenever data information was unavailable for the base year, the prior year has been considered.

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Induced Pluripotent Stem Cells Market Report Research ...

Methods of Producing Thymic Emigrants from Induced …

Induced Pluripotent Stem Cells | Posted by admin
Dec 24 2018

Hematopoietic and pluripotent stem cells can be differentiated into T cells with potential clinical utility. Current approaches for in vitro T cell production rely on Notch signaling and artificial mimicry of thymic selection. However, these approaches result in unconventional or phenotypically aberrant T cells; which may lead to unpredictable behavior in clinical use. Thus, there exists a need for improved methods of generating conventional T cells in vitro from stem cells. Researchers at the National Cancer Institute (NCI) have developed a novel method for the in vitro differentiation of induced pluripotent stem cells (iPSCs) into induced pluripotent stem cell thymic emigrant (iTE) cells. Cells produced by this method are functionally equivalent to natural nave CD8+ T cells. The approach utilizes improved fetal thymic organ culture methodology to mature progenitor cells into conventional T cells in the presence of extrinsic Notch signaling. Cell culture conditions further provide for positive and negative selection to ensure proper maturation. This method produces conventional T cells that are suitable for clinical applications such adoptive cell therapy.

The NCI, Surgery Branch, is seeking statements of capability or interest from parties interested in licensing or collaborative research to further develop, evaluate, or commercialize this method of generating nave T cells from iPSCs.

Image: A visual allegory of the generation of iPSC-derived thymic emigrants (in red). Photo credit: Michael J. Kruhlak, Experimental Transplantation and Immunology Branch (ETIB)

CCR News: https://ccr.cancer.gov/news/article/stem-cell-technology-rejuvenates-can...

Development Stage: Pre-clinical (in vivo)

Related Invention(s): E-133-2017

Inventors:

Raul Vizcardo (NCI)more inventions...

Nicholas Klemen (NCI)more inventions...

Nicholas Restifo (NCI)more inventions...

Intellectual Property: Application No. 62/433,591 Application No. PCT/US2017/065986 Publications:Vizcardo, R, et al. Generation of tumor antigen-specific iPSC-derived thymic emigrants using a 3D thymic culture system. PMID 29562175 Collaboration Opportunity: Licensing and research collaboration

Licensing Contact: John Hewes, Ph.D. Email: John.Hewes@nih.gov Phone: 240-276-5515

OTT Reference No: E-250-2016 Updated: Dec 14, 2018

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Methods of Producing Thymic Emigrants from Induced ...

Global Induced Pluripotent Stem Cells Market Analysis (2018 …

Induced Pluripotent Stem Cells | Posted by admin
Dec 24 2018

The Induced Pluripotent Stem Cells Market (2018 2023): Global Industry Analysis research publication offers readers with a comprehensive knowledge of the induced pluripotent stem cells market scenario in forthcoming years. This report guides through various segments of the global induced pluripotent stem cells market with market size status and forecast 2023. These segments are determined by sizing the market with induced pluripotent stem cells type, end-use segment, and geography. Furthermore, the report offers strategic perspectives on market growth factors such as drivers, restraints, induced pluripotent stem cells market demand and supplier opportunities, technological developments and how they will shape the induced pluripotent stem cells industry.

Market Summary: The main objective of the report is to track the market events such as product launches, induced pluripotent stem cells market ups and downs in terms of volume US$ (mn) and volume (units) from 2013 to 2023, various development activities related to induced pluripotent stem cells products, latest trends, and technologies used in this field. The first overview section of the report comprised with a definition of the global induced pluripotent stem cells market, classification and regional outlook of the market. The regional analysis being used in this report that specifies opportunities available and growth prospects of the global induced pluripotent stem cells market within the specified regions. It additionally provides information related to value chain with a curated list of raw materials suppliers, distributors, induced pluripotent stem cells manufacturers, technological solutions providers and end users of the induced pluripotent stem cells.

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Global Induced Pluripotent Stem Cells Market: Competitive Insights

The crucial section of the report describes the vendor landscape of the global induced pluripotent stem cells market, it includes the profile of leading market players currently operating in the market. The analysis provides information about their market revenues, products manufactured by them, induced pluripotent stem cells manufacturing process and plants, opportunities that are motivating these players and business strategies followed by them. The induced pluripotent stem cells report helps businesses compete better using this scale of reference, although planning their future developments to counter the movements of the other players and stay ahead in the competition.

List of Market Players Profiled in the Report

Segments Covered in the Global Induced Pluripotent Stem Cells Market Report

The research study examines forecasts revenue growth of induced pluripotent stem cells market at global, regional & country levels and provides inclusive insight on the market developments and opportunities available in various segments of the induced pluripotent stem cells market from 2013 to 2023. For the purpose of this study, report segmented the global market based on region, end-user, and induced pluripotent stem cells type. The market shares contributed by these segments are formulated to give the readers a 360-degree assessment of the global induced pluripotent stem cells market.

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What will you discover from induced pluripotent stem cells report?

A comprehensive analysis of current and future market demand for the induced pluripotent stem cells, covering six world regions, end-use industries, growing markets for the induced pluripotent stem cells.

The report employs a combination of primary and secondary research methods for segmenting and estimating quantitative facets of the global induced pluripotent stem cells market.

Exclusive research on established and emerging market players to get competitive advantage of the global induced pluripotent stem cells market.

Extensive analysis of the market drivers, restraints, review of latest trends and technologies used, market openings for the induced pluripotent stem cells.

Details of induced pluripotent stem cells market sizes and five-year forecasts, segmented by product type, end use segment, and region and country worldwide.

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