microbiologist-1332376

Good Tissue Bank Standards of Practice

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What helps make a tissue bank good?

Following Standards of Practice (SOPs) in tissue banking is vital to ensure that every step of human tissue procurement, preparation, and distribution is held to the highest standards. This is done to ensure the safety, quality, and traceability of tissues for surgical use.

The internal Standards of Practice for tissue banks include documents that cover every aspect of this process. These standards cover topics including monitoring tissue recovery sites, the audit of tissue recovery partners, necessary body cooling protocols, microbiological surveillance programs, providing service to tissue donor families, prevention of contamination and cross-contamination at recovery, and many more.

However, the most relevant guide to understanding what makes a “good tissue bank” are the factors that govern “Good Tissue Practice,” defined by the steps that the FDA defines in the manufacturing of human allografts for surgical use: recovery, donor screening and testing, packaging, labeling, distribution, processing, and storage.1 Detailed guidelines for these areas of “Good Tissue Practice” have been developed by the national organization which audits and accredits participating US tissue banks.2

Recovery of Allografts

Recovery is the process of obtaining from a donor either cells or tissues that are intended for human use via implantation, transplantation, infusion, or transfer.1 It is the responsibility of the donor recovery agency to have a program in place that ensures donor information is received, investigated, evaluated, documented, and shared with all agencies that receive tissues from the donor to ensure donor suitability. There are specific SOPs in place to ensure that all recovery personnel are well trained and competent.

There are very specific steps that are taken by the donor recovery operation to control contamination and cross-contamination during retrieval. These steps include adherence to appropriate donor eligibility guidelines such as specific body cooling parameters and time limits for retrieval; ensuring a suitable location for recovery site activities; using clean techniques appropriate to the specific cells/tissues being recovered; cleaning, disinfecting, and sterilization of equipment, supplies, and instruments; monitoring recovery activities for microorganism contamination, such as by culture results; and sharing of all records related to donor eligibility determinations.

Donor Screening and Testing

The main step to prevent potential infectious diseases in the donor tissue supply is primary prevention, which comes from donor screening and testing, first in the form of the detailed medical-social interview (Donor Risk Assessment) of the donor and exhaustive review of their medical records and sexual history. There are multiple levels of review of every record, including by an independent medical doctor specializing in cytopathology. All tissues are also required to be tested by a range of FDA-licensed, cleared, or approved donor screening tests, as applicable by tissue type, such as for HIV, HBV, HCV, syphilis, HTLV, CMV, etc.3 Testing laboratories must also be registered as tissue establishments by the FDA, and are subject to the same rigorous oversight as tissue banks and procurement organizations.

Processing and Storage

Processing is defined by the FDA as “any activity performed on an HCT/P, other than recovery, donor screening, donor testing, storage, labeling, packaging, or distribution, such as testing for microorganisms, preparation, sterilization, steps to inactivate or remove adventitious agents, preservation for storage, and removal from storage.”4 Generally, processing includes cutting, grinding, shaping, culturing, enzymatic digestion, and decellularization of recovered tissues to make them suitable for surgical use.3 Processing must be conducted in a suitable clean room environment with appropriate environmental controls and monitoring.

Processing may also include a sterilization step such as gamma or e-beam radiation. Both are validated, controlled methods of improving allograft tissue safety. Most tissue banks employ a sterilization step in the face of infectious disease risks. To learn more about sterilization through radiation, see this post: “Understanding Tissue Sterilization, Part 2: Radiation Round-up.”

Storage of allografts must be maintained at appropriate temperatures within each tissue’s tolerance limits. A temperature monitoring system must be in place to document storage temperatures and alert staff of deviations before straying outside of acceptable limits. Each unit of stored tissue should be packaged to facilitate sterile storage and prevent contamination or cross-contamination.

Packaging and Labeling

Packaging and shipping are not part of processing, and are therefore not required to be validated or verified in the same way as manufacturing. However, appropriate procedures must be in place to ensure the tissues are protected from contamination and cross-contamination, and that appropriate storage parameters are maintained during transport, particularly that temperature ranges are kept within acceptable limits.

Tissue labeling must make appropriate claims that are supported by verification and validation data to ensure that they are true and accurate. This could include information on sterilization procedures. Labeling also must include a method of internal donor identification, usually a lot-based identifying number on the label to allow for tracking and traceability of every tissue.

Distribution of Allografts

Strict regulations govern the transport and distribution of allografts, including an expiration date for their use. The expiration date is the maximum allowable storage period for the tissue, and expired tissues should not be transplanted. It is also critical to ensuring a safe allograft supply that the distribution of tissues be completely traceable from the original donor, to the retrieval agency, to the intermediary or processor, and on to any other tissue manufacturer or distributors to the ultimate end-user. The information that should travel with an allograft includes tissue ID numbers, tissue type, quantity, time of transport and delivery, recipient of delivery, and who transported and accepted tissues.

If you need help developing a tracking process for your surgical facility, there are resources available to guide you. In addition to the information above, a tissue tracking log should reflect the date of implantation and recipient of every graft. More information on tracking requirements can be found at the FDAs website.

 

How could COVID-19 impact allograft safety?

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It is well known that viruses can sequester in organs and tissues, creating the possibility of transmission during transplant. Until more is known about COVID-19 transmission and sequestration, it’s best to presume that there could be potential risks to patients receiving allografts. Therefore, implementation of guidelines which restrict tissue donation are necessary to preserve the safety of the nation’s allograft supply.

Fortunately, the Physician’s Council of the American Association of Tissue Banks has issued guidance on this matter to the US tissue banking community. Per Dr. Roman Hitchev of the Association, here is a summary of those guidelines:

  • Travel within the last 28 days prior to donation to an area designated by the CDC as Alert Level 2 or Warning Level 3 may constitute grounds for exclusion of a deceased donor regardless of symptoms, or deferral of a living donor for as much time is necessary to ensure at least 28 consecutive days without symptoms1 following the last travel date to the designated area.
  • Fever with severe acute lower respiratory illness (e.g., pneumonia, ARDS) requiring hospitalization without alternative explanatory diagnosis (e.g., influenza) and without a negative SARS-CoV-2 diagnostic test, may constitute grounds for exclusion of a deceased donor, or deferral of a living donor for as much time is necessary to ensure at least 28 consecutive days without these syndromic symptoms.
  • Close contact3 within the last 28 days prior to donation with a person who has confirmed COVID-19 infection or with a Person under Investigation2 (PUI) as defined by the CDC may constitute grounds for exclusion of a deceased donor, or a deferral of a living donor for as much time is necessary to ensure at least 28 consecutive days without symptoms1 following the contact.
  • Confirmed infection or designation of a Person under Investigation2(PUI) as defined by the CDC within the last 28 days prior to donation may constitute grounds for exclusion of a deceased donor or deferral of a living donor for as much time is necessary to ensure at least 28 consecutive days without symptoms after the PUI status is lifted.  

 

  1. Symptoms: Refer to the CDC website – https://www.cdc.gov/coronavirus/2019-ncov/about/symptoms.html
  2. Person under Investigation(PUI): Refer to CDC Criteria to Guide Evaluation of PUI for COVID-19 – https://www.cdc.gov/coronavirus/2019-ncov/hcp/clinical-criteria.html
  3. Close Contact: Refer to the CDC website – https://www.cdc.gov/coronavirus/2019-ncov/faq.html

 

Due to the dynamically changing outbreak geography, medical directors are advised to refer to the CDC on countries at risk for transmission and community spread and the latest travel advisories.

Our country’s tissue banking community is committed to patient safety. The Association and member banks are engaged in research to answer key questions about the new coronavirus and its effects on donated tissues. Allografts.com will be providing updates to the surgical community on the scientific knowledge base as it evolves.

Organs are Allografts, Too!

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Did you know that organs are also a type of allograft? We usually think of skin, tendons, bone, and heart valves as allografts, but solid organs are also allografts, which are defined as “a tissue or organ obtained from one member of a species and grafted to a genetically dissimilar member of the same species.”1

Deceased organ donors can donate two kidneys, one liver, two lungs, one heart, one pancreas, and their intestines. Living organ donors can donate one kidney, one lung, or a portion of their liver, pancreas, or intestine.2 Approximately 115,000 people, including children, await organ transplants in the United States right now. 82% of individuals waiting are waiting for a kidney. There are 8,000 deaths each year in the U.S. because donated organs are not procured in time. 683,000 transplants have taken place in the U.S. since 1988.3

To qualify as an organ donor, blood flow and oxygen must have not been interrupted to organs before tissue recovery, to ensure organ viability. As a result, organ donation requires an individual die under circumstances resulting in a fatal brain injury; usually a massive trauma to the head results in bleeding, swelling, or lack of oxygen to the brain which leads to a lack of brain or brainstem activity and brain death. After all efforts to save the patient’s life have been exhausted, a search is conducted to determine if the patient has personally authorized donation or, if not found on the registry, the next of kin or authorized representative is contacted to be offered the opportunity to authorize donation. Once authorization is obtained, medical and social history is collected from the family. Donation and transplantation professionals determine, based on this information, which organs can be donated and match those organs to patients on the national transplant waiting list.4

In 2014, hands and faces were added to the organ transplant list to be collected from deceased donors.2 The first modern hand transplant was performed in France in 1998. The first hand transplant in the United States occurred one year later, in 1999 in Louisville.5 Less than 100 people have received hand transplants in the world, and less than 30 have received hand transplants in the U.S., largely because of concerns about the risk of rejection, the risks of long-term immunosuppression, and the high level of function that can be obtained with modern prosthetics.6  Hand transplants are still considered experimental by most insurance companies because of a lack of evidence as to their effectiveness; the first hand transplant recipient in France stopped taking his immunosuppressants within two years and requested his transplanted hand be removed, while the first American with a double hand transplant complains that he “can do absolutely nothing” with his transplanted hands, and explored the possibility of getting them removed.5,6

A face transplant is a procedure that replaces all or part of a patient’s face with that of a donor. This is usually performed on individuals who have suffered extreme disfigurement to the face such as through burns, trauma, birth defects, or diseases. Those with severe disfigurement are often ostracized by society for not looking “normal,” and, as the tissue is delicate, they may also have issues with eating and talking. The first face transplant was performed in France in 2005. In 2007, the first full-face transplant was conducted, also in France. The first near-total face transplant was performed in the United States in 2008 at the Cleveland Clinic, which was also the first face transplant to be conducted in the U.S.7 Faces are at higher risk for rejection than solid-organ transplants, since they involve so many different kinds of tissue including muscles, nerves, blood vessels, bones, and skin. Preventing rejection requires a lifetime of taking high doses of immunosuppressive drugs, which makes the patient vulnerable to infections and diseases, especially lymphomas and other cancers.8 The first face transplant recipient, Isabelle Dinoire, developed two types of cancer as a result of prolonged use of immunosuppressants and anti-rejection drugs, eventually leading to her death in 2016.9 Because of concerns about the risks to the procedure, and its relative newness, it is still considered an experimental procedure. Face transplants have also been conducted in several other countries, including Spain, Turkey, China, and Poland.9

A newer, amazing application of allografts are penis transplants, which show incredible promise for restoring urinary function as well as, hopefully, sexual functioning. South Africa has successfully conducted two penis transplants from deceased donors.10 In 2016, Massachusetts General Hospital conducted the first penis transplant in the United States utilizing a genitourinary vascularized composite allograft (GUVCA)—which means it included arteries, veins, nerves, urethra, and skin graft pedicle to properly restore appearance and eventual function.11 Johns Hopkins School of Medicine in Baltimore conducted the first total penis and scrotum transplant in the world in 2018. Utilizing tissue from a deceased donor, they transplanted skin, muscles and tendons, nerves, bone, and blood vessels to repair an injury the patient received serving as a soldier in Afghanistan from an IED.12

As transplant medicine progresses, we may see more successful outcomes with these more complex and challenging types of transplants, which promise to improve the quality of life of many more people.

References

  1. Random House. (2018). Allograft. Dictionary.com. Retrieved from https://www.dictionary.com/browse/allograft
  2. U.S. Department of Health & Human Services. (2018). What can be donated. Organdonor.gov. Retrieved from https://www.organdonor.gov/about/what.html
  3. Donate Life America. (2018). Organ, eye and tissue donation statistics. Retrieved from https://www.donatelife.net/statistics/
  4. Donate Life America. (2018). Deceased donation. Retrieved from https://www.donatelife.net/types-of-donation/deceased-donation/
  5. Skelly, L. M. (2011, March 28). Rare hand transplant surgery successfully performed at Emory University Hospital. Emory University. Retrieved from http://shared.web.emory.edu/whsc/news/releases/2011/03/rare-hand-transplant-surgery.html
  6. Duke Health. (n.d.). Hand transplant: Vascular composite allotransplantation. Retrieved from https://www.dukehealth.org/treatments/transplant-program/hand-transplant
  7. NYU Langone Health. (n.d.). Facial transplant: Eye of the beholder? High School Bioethics. Retrieved from https://med.nyu.edu/highschoolbioethics/briefs/facial-transplantation-eye-beholder
  8. Connors, J. (2018, September). How a transplanted face transformed Katie Stubblefield’s life. National Geographic. Retrieved from https://www.nationalgeographic.com/magazine/2018/09/face-transplant-katie-stubblefield-story-identity-surgery-science/
  9. BBC. (2016, September 6). First face transplant patient Isabelle Dinoire dies in France. BBC News. Retrieved from https://www.bbc.com/news/world-europe-37290986
  10. Akwei, I. (2017, May 23). World’s third penis transplant successfully done in South Africa. Africa News. Retrieved from http://www.africanews.com/2017/05/23/world-s-third-penis-transplant-successfully-done-in-south-africa/
  11. Brown, N. (2016, May 16). First Genitourinary Vascularized Composite Allograft (penile) transplant in the nation performed at Massachusetts General Hospital. Massachusetts General Hospital. Retrieved from https://www.massgeneral.org/News/pressrelease.aspx?id=1937
  12. Kooser, A. (2018, April 23). Wounded US soldier gets first penis and scrotum transplant. CNET. Retrieved from https://www.cnet.com/news/soldier-gets-first-penis-and-scrotum-transplant-johns-hopkins/

 

What is CRISPR? A Primer

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CRISPR is known as a cutting-edge tool for gene editing. Would you be surprised to learn that  that CRISPR originally evolved within single-celled organisms such as bacteria, as their own immune response to viruses? When viruses attack a bacterial cell, they inject their genome—their own viral DNA—into the cell. The bacterial cell, in turn, responds by deploying CRISPR: a scissor-like enzyme called CRISPR-associated protein, or Cas, hooked to a strand of RNA.3 The RNA recognizes the viral DNA, and Cas completely inactivates the targeted gene by making a precise cut in the gene, disabling the viral attack.1

CRISPR is an acronym for “clustered regularly interspaced short palindromic repeats,” which refers to the unique gene organization of bacteria and other microorganisms with short, partially palindromic repeated DNA sequences.2  Since its discovery in bacteria in 2012, scientists have been working in the lab to reproduce the CRISPR method to target plant or animal DNA, rather than viral DNA. CRISPR has the potential to “knock out” specific, targeted genes in plant, animal, or even human cells and, if a strand of DNA coding for a new gene is added, CRISPR can potentially provide a patch in the new gene between the severed ends. If it works, it would actually be fairly simple to design—all a researcher needs is a piece of RNA that can lock on to the targeted gene, attach it to a Cas enzyme, like Cas9, and you have a precision DNA editing tool.1

CRISPR editing technology has the greatest medical potential in humans in utero, to edit genes before birth and prevent the transmission of deadly genetic diseases like Tay-Sachs. Once a person is born with a disease like Tay-Sachs or Huntington’s disease, it’s much more difficult to use CRISPR to treat it.3 Gene editing is also being tested to treat conditions like cancer, blindness, sickle cell, thalassemia, and liver disease—among others.3,4

CRISPR has potentially broad applications that could impact allografts and human tissue engineering. It’s already being used to edit the genomes of mosquitoes, in the hopes of being able to prevent their ability to carry infectious diseases like Zika.3 And CRISPR/Cas is being applied directly within the field of tissue engineering, where it has been used to modify the expression of bone morphogenetic proteins (BMP) improve the osteogenic potential of adipose-derived stem cells (ASC).5

Another application of CRISPR is to edit induced pluripotent stem cell (iPSC) genomes to create special cell lines for use in tissue engineering.5 As techniques are developed to aid in homing and differentiation of stem cells, these customized iPSCs  may allow for the creation of multi-layered tissue scaffolds; cardiovascular grafts, whole organs, and other complex tissues could then be created.

For now, however, medical applications are confined to laboratories.1 Two recent studies have raised concerns that edited cells, designed to treat disease, might trigger cancer, demonstrating the need for further research and refinement before the technique is deployed. Experts describe the cancer risk as “plausible,” but not a “deal-breaker.”4  Considering the potential benefits to humankind, we’ll be staying tuned as the research evolves.

References

[1] Richter, V. (2016, 18 April). What is CRISPR and what does it mean for genetics? COSMOS: The science of everything. Retrieved from: https://cosmosmagazine.com/biology/what-crispr-and-what-does-it-mean-genetics

[2] Pak, E. (2014). CRISPR: A game-changing genetic engineering technique. Harvard: Science in the News. Retrieved from: http://sitn.hms.harvard.edu/flash/2014/crispr-a-game-changing-genetic-engineering-technique/

[3] Delviscio, J. (2018, April 4). How CRISPR works, explained in two minutes. STAT. Retrieved from: https://www.statnews.com/2018/04/04/how-crispr-works-visualized/

[4] Begley, Sharon. (2018, June 12). CRISPR-Edited cells linked to cancer risk in 2 studies. Scientific American. Retrieved from: https://www.scientificamerican.com/article/crispr-edited-cells-linked-to-cancer-risk-in-2-studies/

[5] Pulgarin, D. A. V., Nyber, W. A., Espiinosa, A. (2017, July). CRISPR/Cas systems in tissue engineering: A succinct overview of current use and future opportunities. Current Trends in Biomedical Engineering and Biosciences, 5(4), 001-004. doi:10.19080/CTBEB.2017.05.555670

First Commercial Induced Pluripotent Stem Cell Plant Opens in Japan

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The first commercial facility to produce induced pluripotent stem cells (iPSCs) opened on March 22, 2018. The $340 million, 30,000 square foot facility was built by Sumitomo Dainippon Pharma and is designed to produce stem cells for therapeutic applications.1  Sumitomo Dainippon is currently involved in using iPSCs to create treatments for conditions including macular degeneration, Parkinson’s disease, retinitis pigmentosa, and spinal cord injuries. The plant’s function, initially, will be to produce stem cells for use in clinical trials and early-stage commercial production.2 The more than 30,000 square feet of floor space is divided in three autonomous zones, each producing different types of stem cells. The iPS cell lines come from those produced by Kyoto University’s Center for iPS Cell Research and Application (CiRA).1

The Sumitomo Dainippon Manufacturing Plant for Regenerative Medicine & Cell Therapy is the only facility designed for commercial purposes that exists worldwide. Existing facilities, including the National Institutes of Health’s Center for Regenerative Medicine and the UK Stem Cell Bank, have made stem cells available to researchers, but to date no facility has provided stem cells for commercial purposes. This demonstrates what Masayo Tada, the president of Sumitomo Dainippon, claims is an “overwhelming advantage in the market of medical and pharmaceutical products” that will help the company “occupy a definite position in the field of regenerative medicine.” Sumitomo Dainippon intends to make regenerative medicine one of its core business strategies, and has plans for significant market growth in the coming decade.1

References

  1. Daley, J. (2018, March 23). World’s first commercial iPSC cell plant opens in Japan. The Scientist. Retrieved from: https://www.the-scientist.com/the-nutshell/worlds-first-commercial-ipsc-cell-plant-opens-in-japan-29915
  2. Sumitomo Dainippon Pharma. (2018, March 1). Sumitomo Dainippon Pharma completes manufacturing plant for regenerative medicine and cell therapy. IR News. Retrieved from: https://www.ds-pharma.com/ir/news/2018/20180301-3.html
  3. Photo: Immunocytochemistry of Kindey Tubular Epithelial Cells derived from induced pluripotent stem cells. By Ugnezija7, from Wikimedia Commons

 

Understanding Stem Cells Part 2: Ready for the Clinic?

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With advances in research and technology, using stem cells for their regenerative potential has become a reality. But many stem cell treatments on the market have been marketed to the public and applied clinically without scientific evidence of efficacy. Worse, some treatments with stem cells have caused serious adverse events.

Provided that cell and tissue products fall into the “minimally manipulated” classification established by the Food and Drug Administration (FDA) under Title 21 of the Code of Federal Regulations Part 1271, studies to prove efficacy and safety are not required. This makes it possible to promote stem cell treatments for any use, without scientific or medical evidence that they work. Medical practitioners and patients alike must be aware that stem cell treatments are generally not “FDA approved.”

Draft guidance for industry was issued by FDA in 2017, proposing guidelines for evaluating and approving regenerative therapies which have demonstrated safe and efficacious use in serious conditions. These products could achieve a designation of “Regenerative Medicine Advanced Therapy” or “RMAT” in several ways. While the actual policy is still being formulated, this guidance initiated discussion with industry stakeholders and signaled increasing attention by FDA to regulating products touted as “regenerative,” including stem cells.

Cells for stem cell treatments are often autologous, extracted from the patient through bone marrow aspiration, a blood draw and centrifuge process, or liposuction (for adipose stem cells). There are also stem cell products available commercially that are allografts (cells or tissue from a human donor), including umbilical cord blood stem cells, allograft fat, and cell-based bone grafts. The number and type of stem cell varies based on the tissue of origin and the processing of the product, and both of these factors can be expected to affect outcomes.

Stem cell clinics often charge very high prices for stem cell treatments, from $1,500 for a joint injection to $25,000 for a systemic treatment. Insurance typically does not cover these treatments because they are considered experimental. Patients range from injured athletes and weekend warriors who want to avoid surgery, to desperate and terminally ill patients who are struggling with incurable conditions.

One unfortunate example of stem cell treatment gone wrong occurred at a Florida clinic. Three elderly women received stem cell injections to treat age-related macular degeneration and were blinded by the procedure. One of the women was apparently misled into believing that she was taking part in a clinical trial. The clinic which performed the injections, Bioheart Inc., had not conducted any pre-clinical or clinical trials. Today, Bioheart Inc. is doing business as US Stem Cell, which received a warning letter from FDA in August 2017, citing a multitude of deviations from current good manufacturing processes as well as inappropriate marketing claims and clinical uses. Read about it here.

Yet, there are many reasons to be excited about stem cells which are produced, processed, and applied appropriately. The California Institute for Regenerative Medicine (CIRM) has pioneered stem cell treatments to cure children afflicted with Severe Combined Immunodeficiency Disease (SCID), and they are now entering Phase 2 clinical trials for this clinical application. CIRM has over 40 trials running at this time, all targeting devastating medical problems.

In more common clinical scenarios, studies have shown that stem cell injections can improve osteoarthritis symptoms, and many sports medicine and orthopedic specialists are using them to help patients avoid surgery.

Future cardiac applications may become possible, too. In an article published in Nature Communications by Shadrin and colleagues (2017), scientific work investigated the development of a scaffold composed of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), which are heart cells derived from adult human stem cells. The scientists developed a “cardiopatch”, essentially a network of organized heart cell tissue, and conducted experiments in a rodent model. The cardiopatches became vascularized, showed specific characteristics of adult heart tissue, and conducted the heart’s electrical charge. The researchers believe that their 4x4cm cardiopatches could eventually be scaled to create larger sections of functioning tissue for myocardial repair.

Validated treatments using stem cells have the potential to improve countless lives. But, for most conditions, we are years away from establishing proper protocols and doses. It’s appropriate for practitioner and patient alike to weigh the risks and benefits of any treatment, including (and, perhaps, especially) those promising stunning disease reversals and health advancements. Meanwhile, research continues to bring the promise of stem cells closer into focus and, step by step, safely into the clinic.

 

References

Begley, Sharon. (2017, March 15). Three patients blinded by stem cell procedure, physicians say. STAT. Retrieved from https://www.statnews.com/2017/03/15/stem-cell-patients-blind-macular-degeneration/.

Doheny, Kathleen. (2017, April 14). Stem cells for knees: promising treatment or hoax? WebMD. Retrieved from https://www.webmd.com/osteoarthritis/news/20170407/stem-cells-for-knees-promising-treatment-or-hoax#1.

GEN News Highlights. (2018, January 29). Stem cells: On your mark, get set, don’t go. Genetic Engineering & Biotechnology News. Retrieved from https://www.genengnews.com/gen-news-highlights/stem-cells-on-your-mark-get-set-dont-go/81255432.

McFarling, Usha Lee. (2016, February 8). FDA moves to crack down on unproven stem cell therapies. STAT. Retrieved from https://www.statnews.com/2016/02/08/fda-crackdown-stem-cell-clinics/.

Shadrin, Ilya Y., Allen, Brian W., Qian, Ying, Jackman, Christopher P., Carlson, Aaron L., Juhas, Mark E., & Bursac, Nenad. (2017, November 28). Cardiopatch platform enables maturation and scale-up of human pluripotent stem cell-derived engineered heart tissues. Nature Communications, 8: 1825. doi:10.1038/s41467-017-01946-x

Uygur, Aysu, & Lee, Richard T. (2016, February 22). Mechanisms of cardiac regeneration. Developmental Cell, 36: 362-374. https://doi.org/10.1016/j.devcel.2016.01.018

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Understanding Stem Cells, Part 1: Stem Cells 101

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Stem cells are a promising regenerative medicine tool and treatment. So much information on stem cell research exists that sorting through the facts and the hype can be daunting. This post will focus on explaining stem cell basics: What are stem cells, and what are the different types? Here, we will focus on adult stem cells, also known as somatic stem cells, since these are most relevant to medical research and clinical use today.

Nearly every cell in the body has a specific purpose. When a fetus begins to develop, cells become specialized (or differentiated), developing structure and characteristics specific to carrying out a special function in the body.1 For example, heart muscle cells generate movement (i.e., heartbeat), and red blood cells contain hemoglobin so that they can transport oxygen throughout the bloodstream. Stem cells, on the other hand, are unspecialized (undifferentiated); they have the potential to become any type of cell.

There are different categories of stem cells. Most current research focuses on these stem cell types:

  • Totipotent—can form all cell types in the body. During embryonic development, totipotent stem cells differentiate into all of the different cell types in order to form a whole organism. Researchers have been able to take differentiated cells and return them to a state of totipotency. More work is needed to make totipotent cells clinically useful.
  • Pluripotent—are descended from totipotent stem cells and can form into almost all cell types. Pluripotent stem cells can form any type of cell derived from the three germ layers: endoderm (forms the inside lining of stomach, digestive tract, and lungs), mesoderm (forms blood, muscle, and bone), and ectoderm (forms the skin and nervous system.) Pluripotent stem cells naturally exist in the embryo.
  • Multipotent—more specialized than totipotent and pluripotent stem cells, multipotent cells can develop into several different somatic cell types. Multipotent stem cells give rise to various tissues and organs during development, and some of these stem cells remain in a state of dormancy (“quiescence”) within their specific tissue niches of the adult human body. Types of multipotent stem cells include:
    • Hematopoietic stem cells—form the components of the blood.
    • Mesenchymal stem cells—form bone, cartilage, muscle, and fat cells.
    • Neural stem cells—form the nervous system.
    • Epithelial stem cells—form cells in the digestive tract lining.
  • Oligopotent—can form a few different cell types. They are derived from multipotent stem cells, descending from hematopoietic stem cells. Examples:
    • Lymphoid cells—form several types of immune cells, such as B cells and T cells.
    • Myeloid cells—form red blood cells and white blood cells, among others.

As mentioned above, stem cells are found in the adult body within tissues such as blood, bone, skin, heart, and gut, as multipotent cells. It’s believed that the cells are located in a stem cell niche, or a specific area of each tissue. The number of niche-dwelling stem cells is relatively small. These cells may remain quiescent (undifferentiated) for a long time, until disease, injury, or normal body processes cause the need for differentiation.

Three main sources of these adult somatic stem cells are bone marrow, blood, and adipose, tissue. Research is being conducted on ways to grow larger numbers of adult stem cells outside the body. When experimental conditions in the laboratory are right, stem cells can be induced to become specialized cells that perform specific functions. This has important implications for regenerative medicine.

Inducing cells from a patient’s own body to grow into the specific tissue they need to receive could lead to tremendous medical advances, someday eliminating the need for allografts and organ transplants. In the lab, researchers have been able to manipulate adult somatic stem cells to become induced pluripotent stem cells, or iPSCs. Through forced expression of specific genes, transcription factors, and epigenetic factors, differentiated somatic cells can be de-differentiated or induced to return to a pluripotent state. Advances have been made in scientific research on therapeutic implications of iPSCs.

Another source of pluripotent stem cells is umbilical cord blood. Stem cells from umbilical cord blood can be saved just after birth, cryopreserved for later use, and stored in an umbilical cord blood bank.2 There are now commercialized injectable therapies using umbilical cord blood stem cells.

There are many exciting possible applications of the stem cell research now being performed in laboratories worldwide. One example of this involves limbal stem cells, or LSCs. LSCs are found in the limbus, the part of the eye where the cornea meets the sclera. Normally, these cells are used to regenerate the cornea, which has a very fast turnover rate of 1-2 weeks in the human body. If a person’s LSCs deteriorate or they do not have enough LSCs, their corneas degenerate and they will go blind. However, research suggests that if LSCs were transplanted into such an eye, the entire cornea would regenerate.3 Researchers have studied LSCs using experiments with mice, and were able to restore the cornea of mice with experimental blindness. These results are promising for regenerative medicine in humans. For more information about the applications of stem cell research, check out Part 2.

References:

Stem Cell Basics (2016). National Institutes of Health. Retrieved from https://stemcells.nih.gov/info/basics.htm.

Jawdat, Dunia (2016). Banking of Human Umbilical Cord Blood Stem Cells and Their Clinical Applications. In Abdelalim E. (ed.) Recent Advances in Stem Cells. Humana Press. Retrieved from https://link.springer.com/chapter/10.1007/978-3-319-33270-3_8.

Limbal Stem Cells for Treatment of Corneal Blindness (2017). Advanced Science News. Retrieved from http://www.advancedsciencenews.com/limbal-stem-cells-treatment-corneal-blindness/.

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3D Printed Tissues From Bench to Bedside

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3D bioprinting has the potential to provide significant medical advances. For example, 3D printing of functional tissues and organs could someday alleviate shortages in organ availability and the problem of organ incompatibility. But where does the field stand today?

A look at the research benches of top labs can give us an idea. In this post we’ll take a look at CELLINK’s innovative inks and printers, ORGANOVO’s functional bioprinted tissues, Dr. Anthony Atala’s Wake Forest Institute for Regenerative Medicine, and the 3D printed cortical fibers line by Xtant Medical, 3Demin®.

CELLINK is the world’s first bioink company. A Swedish company founded in 2016, CELLINK solved problems for the tissue engineering community by commercializing hydrogel bioinks which can be used to 3D print cartilage, bone, skin, or muscle. CELLINK is also a top provider of bioprinters. It’s likely to be one of CELLINK’s printers in a researcher’s lab being used to create 3D printed allografts during experiments, as was the case of the 3D printed cartilage mentioned in the prior post. With the right raw materials, it’s possible to create multiple biologics scaffolds and structures, and CELLINK is at the forefront of bioink development.1

ORGANOVO creates functional, responsive bioprinted human tissue models for preclinical testing and drug discovery research. Their 3D bioprinted tissues remain viable and dynamic for more than 28 days in vitro.2 Their key architectural and functional features mimic natural function and provide important information for scientists.

Pharmaceutical companies conducting research can examine the effects of new compounds on ORGANOVO’s ExVive™ 3D bioprinted human tissues. This allows tissue-specific data to be collected to better predict human outcomes, lower total development time, and reduce costs.

For example, liver and kidney tissues can be used to evaluate drug exposure for both acute and chronic toxicity and metabolism studies.2 ORGANOVO is also working on developing functional, 3D tissues for implantation or delivery into the human body, with the intention of creating these tissue for use for repair or replacement of damaged or diseased tissues.3

The Wake Forest Institute for Regenerative Medicine is one of the nation’s top research labs working to translate scientific discoveries in 3D printing into clinical therapies. Led by Dr. Anthony Atala, a pioneer in medical tissue engineering, this interdisciplinary team is engineering more than 30 different replacement tissues and organs to develop curative cell therapies.4 Wake Forest was the first institution to successfully engineer laboratory-grown organs and implant them into humans: these were engineered bladders grown from the patient’s own cells.5

The fact is, all living tissues are composed of many different cell types arranged in a specific order that is essential to function. Therefore, when using 3D printing to create a tissue, precision is critical.6 Researchers at Wake Forest have designed their own inkjet bioprinter that can print kidney cells, and the binders to hold the cells together, into a 3D kidney prototype. This bioprinter could potentially be used to create a custom organ based on a CT scan and patient data. Their bioprinter is also being tested in the creation of other structured tissue, such as the ear.7

While 3D bioprinting of living, functional, clinically applicable tissues and organs remains the Holy Grail, there are much simpler examples of the technology already commercialized and in use today.  Bacterin’s 3Demin® line of demineralized cortical bone fibers is bioprinted into different shapes and sizes designed for specific surgical applications. The bioprinting process creates a porous, interconnected allograft that contains BMPs and other growth factors necessary to promote new bone formation. It is an osteoconductive demineralized cortical fiber matrix with osteoinductive potential that can be used as a stand-alone graft, or in combination with autologous bone, bone marrow, DBM putty, and other products.8

Bacterin’s production of customized bone implants using 3D printing technology was applied clinically during a mission sponsored by the World Craniofacial Foundation. In Zambia, a child named Grace Kabelenga was born with the most severe facial cleft in 40 years (Tessier 0-14), and a total absence of the bones separating her brain from her oral cavity.9 3D printed plastic prototypes allowed the team to simulate surgery and to create molds for custom bone implants. Bacterin’s 3Demin® bone allograft material was used to create custom 3D printed bone to serve as a “bandeau” to define the superior orbital rims above her eyes.10 This was combined with other Bacterin allografts and rhBMP2 to reconstruct missing parts of Grace’s skull.

Multiple surgeries have allowed Grace to function as a normal child would. Allografts.com members can view a video about the WCF mission on our portal. A slide deck is also available here.

3D printing for tissue engineering has the potential to provide incredible benefits to humankind if the technology can be developed to its full potential for medical applications.

If you missed it, you can read more about the history and applications of bioprinting here: 3D Printing in Tissue Engineering.

References:

1 CELLINK. (2017). BioX Printer. Retrieved from https://cellink.com/bioprinter/

2 ORGANOVO. (2017). 3D human liver tissue testing services. Retrieved from http://organovo.com/tissues-services/exvive3d-human-tissue-models-services-research/3d-human-liver-tissue-testing-services/

3 ORGANOVO. (2017). About ORGANOVO. Retrieved from http://organovo.com/about/about-organovo/

4 Wake Forest School of Medicine. (n.d.). Wake Forest Institute for Regenerative Medicine. Retrieved from http://www.wakehealth.edu/WFIRM/

5 Wake Forest Baptist Medical Center. (2006). Wake Forest physician reports first human recipients of laboratory-grown organs. Retrieved from http://www.wakehealth.edu/News-Releases/2006/Wake_Forest_Physician_Reports_First_Human_Recipients_of_Laboratory-Grown_Organs.htm

6 Wake Forest School of Medicine. (2017, September 6). Using 3D printing technology to print organs and tissue. Retrieved from http://www.wakehealth.edu/Research/WFIRM/Our-Story/Inside-the-Lab/Bioprinting.htm

7 Wake Forest School of Medicine. (2016, August 8). Replacement organs and tissues: Engineering a kidney. Retrieved from http://www.wakehealth.edu/Research/WFIRM/Research/Engineering-A-Kidney.htm

8 Bacterin. (2015). 3Demin Benefits. Retrieved from: https://xtantmedical.com/app/uploads/2016/09/3Demin_Brochure_Digital-14009B.pdf

9 3D Systems. (2016). Delivering the gift of a normal life. Retrieved from: https://www.3dsystems.com/sites/default/files/2016/cs_3ds_grace_kabelenga_0316_0.pdf

10 3D Systems. (2017). Delivering the gift of a normal life with 3D systems healthcare solutions. Retrieved from 3D Systems. Delivering the gift of a normal life with 3D systems healthcare solutions. Retrieved from https://www.3dsystems.com/learning-center/case-studies/delivering-gift-normal-life

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Hot Topics: Zika Virus and Allografts, Part 1

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Background

Since the Zika outbreak first began in Brazil in late 2015,1 its spread has been of significant concern to the medical community. First identified due to the significant rise in microcephaly among newborns in Brazil, Zika infection has also been associated with other major birth defects including neural tube defects, eye abnormalities, brain abnormalities, and central nervous system dysfunction unrelated to microcephaly.

Studies have estimated the rate of birth defects among completed pregnancies with Zika infections to be 1 in 20, or 5%, with a higher risk of birth defects when infection occurs earlier in pregnancy.2 In addition to complications during pregnancy, Zika infection also has been shown to cause Guillain-Barré syndrome in adults and may result in other lasting neurological complications.3

Spread in the United States

The Zika virus is spread by two species of mosquito, Aedes albopictus and Aedes aegypti. Both of these vectors are native to the United States, but they are at the northernmost range of their territory and are therefore not found in every state. Ten states do not fall into the estimated range of either vector species and are therefore believed to be free of any native Zika transmission risk. The other states, as seen in the maps below, have at least small areas of potential mosquito transmission, though only Florida and Texas are known to have had local mosquito transmission of Zika to date.

Estimated range of Aedes albopictus and Aedes aegypti in the United States, CDC 2016*
*Maps are not meant to represent risk for spread of disease.

Zika became a nationally notifiable disease in 2016, which mandates reporting of cases to the CDC.4 As a result, there is very good surveillance of the disease in symptomatic patients, though many adults infected with Zika may remain asymptomatic or have only mild, flu-like symptoms.5

In 2016, the United States had 5,102 symptomatic cases of Zika reported to the CDC. Of these, only 224 were acquired through domestic mosquito-borne transmission (218 in Florida and 6 in Texas).4 There is a significantly declining prevalence of Zika virus disease in both North and South America, primarily due to the success of robust mosquito control measures and the public’s attention to preventing exposure.2

As of July 26, 2017, the United States has only had 181 symptomatic cases of Zika reported to the CDC for 2017. None were due to domestic transmission.6 It is believed that all prior areas of local mosquito transmission have been eliminated within the United States.

Clinical Testing for the Zika Virus

No diagnostic tests for the Zika virus are presently approved by the FDA for use in patients, though a number of tests have been developed. These tests are available due to an Emergency Use Authorization (EUA) for use in symptomatic patients residing in (or who have recently traveled to) an area with a risk of Zika infection. A total of 15 different tests are currently available under the Zika EUA, including molecular tests like real-time PCR and serological tests like ELISA. The Trioplex Real-Time RT-PCR assay (Nucleic Acid-based Testing) and the Zika MAC-ELISA (IgM antibody-based testing) are the only two tests that are in widespread use and distribution.7

Tests like these are important and clinically useful, but because of their newness and lack of FDA approval through the typical Premarket Application (PMA) process, concrete data on sensitivity, specificity, and predictive values are still being collected. Testing poses three primary challenges. First, the Zika virus is only transiently present in body fluids, which makes identifying the virus difficult using PCR NAT-based molecular testing. Second, serological testing for IgM antibodies using the MAC-ELISA cannot reliably pinpoint whether infection occurred during or prior to a pregnancy. Finally, serologic tests have been found to have cross-reactivity with other flaviviruses like dengue and are prone to false-positive results, which is a problem in zones where infection rates are high for the other flaviviruses.2

Current guidelines continue to recommend testing of symptomatic pregnant women with recent possible Zika exposure. However, the CDC now only recommends that asymptomatic pregnant women with ongoing possible Zika virus exposure be offered Zika virus NAT testing three times during pregnancy. IgM testing is no longer recommended for routine testing in pregnant women with ongoing Zika exposure, since IgM can persist for months after infection and therefore cannot reliably determine whether an infection occurred during the current pregnancy.2 The CDC also no longer recommends routine screening of asymptomatic pregnant women without ongoing Zika exposure, except in cases where prenatal ultrasound findings are consistent with congenital Zika virus syndrome to assist with establishing a diagnosis for the fetus.2

Testing for Zika in Human Allografts

Right now there is no test available to detect the Zika virus in human allografts, a category which includes placental tissues like the amniotic membrane and umbilical cord. The Procleix Zika Virus Assay is currently being tested under an Investigational New Drug (IND) protocol by manufacturer Gryfols and Hologic in conjunction with industry partners, but studies have not yet been completed to allow for FDA approval. The Procleix assay system, if proven effective, will be able to screen blood, plasma, and organ and tissue donations for the Zika virus.

You can read more about how tissue banks are managing risk for Zika virus in this blog post: Hot Topics: Zika Virus and Allografts, Part 2

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Hot Topics: Zika Virus and Allografts, Part 2

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There is presently no test available to detect the presence of Zika virus in allograft tissues. This may change if the Procleix Zika Virus Assay now being investigated proves to be an accurate and reliable test. Until then, how are tissue banks managing the risks posed by this viral threat?

It’s important to understand that Zika is not the first virus to affect tissue banking, and the industry has specific protocols already established to protect the tissue supply. This begins with what happens before tissue procurement and acceptance, as the key to keeping Zika (and other viruses) out of the supply chain lies in primary prevention.

Tissue banks long ago implemented stringent donor screening methods and medical records review that form the basis of determining high risk for viruses such as HIV and hepatitis. These same screening methods are effective at assessing risk of Zika, along with other new contagious diseases such as West Nile, Dengue, and Chikungunya. Both deceased and living donors are screened for infectious disease risks routinely, although more public attention is focused on Zika at this time.

In the case of screening living donor mothers for Zika, Allografts.com owner Nikki Couloumbis turned to Craig Thomsen, the Director of Quality Systems for TissueTech, the Miami-based market-leading producer of cryopreserved placental tissues provided by Bio-Tissue and Amniox Medical.

Nikki: “Craig, can you discuss what TissueTech is doing to protect the placental tissue supply from Zika?”

Craig: “Absolutely. First, we do not work with any procurement partners that collect tissues within CDC-identified Zika transmission zones, including Miami-Dade, Broward, Palm Beach in Florida, and Brownsville, Texas. So, tissues potentially most at risk of infection are excluded, including the ones right in our backyard.

Second, we follow procedures that would disqualify outright those donors at risk of Zika. The main step to prevent potential infectious diseases has always been the Donor Risk Assessment Interview.”

Nikki: “In the case of deceased donors this interview is conducted with next-of-kin. But the same type of exhaustive review of risks is conducted with living donors. Can you explain some aspects of what you look for?”

Craig: “The DRAI is a very effective tool. It’s extremely in-depth, with very specific questions mandated by Federal and State authorities, supplemented by specific questions guided by company-specific exclusions. The interview addresses social behavior including drug and alcohol use, sexual partners, and tattoos and piercings. It includes a detailed family history, medical history, and travel history for the mother and her partner to known Zika transmission zones. Any living donor who has lived or traveled to an active Zika virus transmission zone within the last 16 months is not eligible to donate.”

“There are multiple levels of review of every record, including by an independent medical doctor specializing in cytopathology, and the exclusion criteria are very stringent, mandated by both the FDA and TissueTech Standards of Practice.”

Nikki: “I expect that in most cases, women at risk are self-identifying as well, so there is less likely to be a case that gets missed.  Furthermore, donors are uncompensated, so there is no incentive to donate if they pose a risk of transmitting Zika. Right?”

Craig: “That’s correct. The public’s high awareness of Zika makes it less likely that a tissue at risk will be donated to begin with. Donors with known infectious disease risks won’t be referred by their obstetricians for tissue donation, and pregnant women living in areas outside of transmission zones who believe they are at risk typically self-identify to their medical providers and request a test. This would be a part of their medical record, and it automatically excludes the mother from birth tissue donation.”

Nikki: “Clearly, it is highly unlikely that Zika-infected tissue will be processed with the extensive procedures that are in place. That’s good news. But let’s discuss your thoughts on sterilization methods. Irradiation has been used by some tissue banks to address concerns about other viruses in allografts.”

Craig: “While that is true, there is no validated method for destroying Zika virus in tissues. The amount of radiation required has not been quantified. So even radiated tissues would pose a risk if primary prevention procedures lapsed. And in the case of placental tissues, biologic value and quality would likely be affected. Truthfully, radiation is not necessary if strict exclusionary procedures are followed absolutely, and that is what we do at TissueTech.”

Nikki: “What is the news on a laboratory test for allografts down the road?”

Craig: “TissueTech has joined with leaders in the industry to jointly petition FDA to amend its recommended guidelines to include testing the donor mother’s blood as a screening tool. The FDA has already approved Zika blood tests for blood donation, so testing placenta donors would seem a logical extension of that. FDA has also approved an investigational study of an assay for Zika virus in tissues which would include placentas. So, yes, we expect progress this year and next year.”

Nikki: “Thanks, Craig, very informative. Our readers will appreciate your insights.”