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 JoVE Biology

Tracheotomy: A Method for Transplantation of Stem Cells to the Lung

1

1Dept. of Cell Biology, Harvard Medical School

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    Aseptic techniques are used th

    Summary

    Significant breakthroughs in stem cell identification are continuously being made. To translate these discoveries, however, novel methods for cellular delivery must be devised. Here I report that the airways provide a safe route for stem cell transplantation to the lungs.

    Date Published: 2/25/2007, Issue 2; doi: 10.3791/163

    Cite this Article

    Peter, Y. Tracheotomy: A Method for Transplantation of Stem Cells to the Lung. J. Vis. Exp. (2), e163, doi:10.3791/163 (2007).

    Abstract

    Lung disease is a leading cause of death and likely to become an epidemic given increases in pollution and smoking worldwide. Advances in stem cell therapy may alleviate many of the symptoms associated with lung disease and induce alveolar repair in adults. Concurrent with the ongoing search for stem cells applicable for human treatment, precise delivery and homing (to the site of disease) must be reassured for successful therapy. Here, I report that stem cells can safely be instilled via the trachea opening a non-stop route to the lung. This method involves a skin incision, caudal insertion of a cannula into and along the tracheal lumen, and injection of a stem cell vehicle mixture into airways of the lung. A broad range of media solutions and stabilizers can be instilled via tracheotomy, resulting in the ability to deliver a wider range of cell types. With alveolar epithelium confining these cells to the lumen, lung expansion and negative pressure during inhalation may also assist in stem cell integration. Tracheal delivery of stem cells, with a quick uptake and the ability to handle a large range of treatments, could accelerate the development of cell-based therapies, opening new avenues for treatment of lung disease.

    Protocol

    Pre-operative

    1. Recipient mouse and donor cells
    2. Operating platform with tape and suture thread
    3. 1cc syringe
    4. 23G needle
    5. Anesthetic: avertin 25 mg/ml (alternatively ketamine 9 mg/ml and xylazin 1 mg/ml)
    6. Hair clipper with #40 blade
    7. Sterile gloves
    8. Alcohol swabs
    9. Iodine
    10. Sterilized surgical instruments
    11. Suture thread
    12. 22G catheter
    13. 200 ul pipette (sterile)
    14. Sterilized pipette tips
    15. Antibiotic glue ointment
    16. Sharpie container

    Operative

    Aseptic techniques are used throughout this procedure.

    1. Weigh mouse.
    2. Intraperitoneal injection of anesthetic (250 mg/kg avertin or alternatively ketamine 90 mg/kg and xylazine 10 mg/kg) at doses sufficient to allow spontaneous breathing, but provide surgical-concentration anesthesia. For surgery to commence the toe pinch was used as an indicator of deep pain with no response from the anesthetized animal.
    3. Lay mouse with incisors tucked in suture thread and tape tail to stabilize.
    4. Clip hair using a #40 clipper blade.
    5. Change gloves (continue with sterile gloves).
    6. Disinfect twice with alcohol prior to skin incision.
    7. Sterilize area with an iodophore.
    8. Using sterile instruments make a small incision (<1 cm) parallel to the trachea.
    9. Carefully separate tissue from cartilage.
    10. A tie is inserted underneath the trachea.
    11. Gently raise tie upward and insert needle into exposed trachea.
    12. Remove needle and insert the tube smoothly, yet rapidly, until feeling slight resistance.
    13. Pipette cells (up to 50 microliters volume) directly into the catheter.
    14. Immediately remove tube and tie while liberating mouse from operating apparatus.
    15. If needed gently hold mouse upright (vertical) by clinging his anterior paws and move gently.
    16. Close the wound with one suture or antibiotic glue.

    Post-operative

    1. Keep mouse warm.
    2. Buprenorphine (0.05 mg/kg) should be given as post-operative analgesia every 12 hours for the first 24 hours following this procedure.

    Figure 1

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    Disclosures

    Materials

    Name Company Catalog Number Comments
    23G needle
    1cc syringe
    22G catheter
    suture thread
    antibiotic glue ointment
    Anesthetic Avertin 25 mg/ml. Alternatively ketamine 9 mg/ml and xylazin 1 mg/ml.

    Comments

    4 Comments

    I agree with the previous comments about the anesthesia. This is one of the first videos I have watched on JOVE and I think the idea of video journalling is great, but there seems to be potentially dangerous animal care issues for the journal and authors since the use of animals is plainly visible as opposed to written descriptions...
    Also this technique should really be performed through intubation with no surgery.

    Reply

    Posted by: AnonymousJanuary 22, 2008, 1:50 PM

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