Exosomes and Dry Eye Disease: its Role in Regenerative Medicine

Exosomes have become a better-understood cellular vehicle or vesicle to help stimulate the body’s regenerative properties.

These little vesicles are extracellular vesicles (EVs) with a diameter of about 40-100nm which are produced in the endosomal compartment of most eukaryotic cells.

They are composed of lipids, proteins, mRNA, and miRNAs which appear to work to stimulate the body’s own progenitor cells.

There is a great deal of research on Exosomes as they seem to be the key component as to why stem cells (ie, Bone Marrow Transplantation), may work to heal cancer. Many studies have indicated that exosomes are key in wound healing and they mediate the regenerative outcomes in tissue injury. Mesenchymal stem cell exosomes were found to activate several signaling pathways important in wound healing, bone fracture repair, immune-mediated responses, and inflammatory diseases. Exosomes induce the expression of a number of growth factors:

1. hepatocyte growth factor (HGF)
2. insulin-like growth factor-1 (IGF1)
3. nerve growth factor (NGF), [this plays a key role in eye pain from dry eye
4. stromal-derived growth factor-1 (SDF1)). 

Scientists have also found that exosomes released from oral keratinocytes can accelerate wound healing, even when human exosomes were applied to rat wounds not corneal or eye wounds but still very interesting. 

MSC exosomes may be useful for dermatologic and ophthalmological applications. Exosomes improve wound healing, by affecting fibroblasts, keratinocytes and endothelial cells. Other in vitro studies have shown the ability to regrow a hair follicle with the regeneration of dermal papilla cells. In vivo studies of MSC exosomes have shown accelerated wound healing, reduction in scarring, modulation of the thermal injury in burns, reduced inflammation in dermatitis,

Exosomes can travel in body without risk of clumping in the lungs and thus many patients have had exosomes transfused into their bloodstream. The research is controversial. Here are 2 sides to the story & more research below.

No one has used Exosomes as an eye drop nor insterted exosomes into meibomian glands. Our hope is to explore this option for safety purposes in the near future with future prospective, randomized, double blinded studies.  


  1. https://kimeralabs.com/kimera-labs-meets-with-medical-advisory-boards-to-develop-dermatology-and-oncology-applications-of-msc-exosomes/
  2. https://en.wikipedia.org/wiki/Exosome_(vesicle)
  3. Elahi FM, Farwell DG, Nolta JA, Anderson JD (August 2019). “Preclinical translation of exosomes derived from mesenchymal stem/stromal cells”Stem Cells0 (1): 15–21. doi:10.1002/stem.3061PMC 7004029PMID 31381842.
  4. ^ Basu J, Ludlow JW (2016). “Exosomes for repair, regeneration and rejuvenation”. Expert Opinion on Biological Therapy16 (4): 489–506. doi:10.1517/14712598.2016.1131976PMID 26817494S2CID 10370397.
  5. ^ “MSC-derived Exosomes Promote Bone Fracture Repair”Stem Cells Portal. 2 January 2017.
  6. ^ Silva AM, Teixeira JH, Almeida MI, Gonçalves RM, Barbosa MA, Santos SG (February 2017). “Extracellular Vesicles: Immunomodulatory messengers in the context of tissue repair/regeneration”. European Journal of Pharmaceutical Sciences98: 86–95. 

Controversy on Exosomes:
Dr. Douglas Spiel   VERSUS  Paul Knoepfler, Ph.D.

Pro Exosomes
Pro Stem Cells     VERSUS    the other side

Dr. Spiel works at Kimera Research & Boston BioLife:

Dr. Spiel seems to be well respected in his field as an interventional pain MD and radiologist and is likely fed up with prescribing steroids and opioids to get rid of chronic pain. It seems this was a motivating factor to begin using stem cells. He has been on the front lines of patients with chronic pain for years.

Dr. Koepfler is a well-respected PhD at UC Davis. He is not an MD so does not treat patients it appears as a daily practice. He is excellent at what he does and has been very careful not to promote stem cell use given the limited data and the absence of any randomized, double-blinded, controlled studies. He has excellent points on his blog.

Paul Knoepfler

The Knoepfler Lab conducts developmental biology, stem cell, and cancer research with a focus on chromatin and epigenetic mechanisms at the UC Davis School of Medicine, Sacramento, CA 95817. We are particularly interested in regulation of normal brain growth and how epigenomic programming goes awry in childhood brain tumors and microcephaly.

You can contact Dr. Knoepfler at: knoepfler@ucdavis.edu.

The Knoepfler Lab currently receives funding from NIH and the Alex’s Lemonade Stand Foundation.

The answer to whether Stem Cell use and/or Exosomes are the wave of the future is still unclear. The majority of MDs view these “autologous” options as generally safe. Is it worth the money? That is the biggest concern right now as these treatments are not covered by insurance. 

More information is located below. My own research has convinced me that many patients are having chronic eye pain with all the available FDA approved options and we have to find better alternatives. We have used autologous serum and PRP drops for years with excellent results. The ability of the body to heal itself should not be underestimated. I think stem cells and exosomes will someday be a standard treatment option (as currently is limbal stem cell transplantation for severe limbal stem cell deficiency) but we need funds to do proper, large scale, double-blinded, randomized controlled studies. 


Comparison of CVs:

1. Douglas Spiel, M.D.
Interventional Pain Specialist and Board Certified Radiologist,
Spiel MD

Dr. Spiel’s unique clinical background has enabled him to view the practice of interventional pain through a broader perspective. By elevating the diagnosis and treatment of those in pain to an art form, he is uniquely qualified to blend the objective and subjective components of a patient’s signs and symptoms to arrive at a plausible and treatable diagnosis.

Dr. Spiel is widely recognized as an expert in numerous treatment modalities and has lectured both throughout the country and abroad. He credits the numerous physicians worldwide that he has studied with as providing the necessary scaffold to build his current diverse skill set. Dr. Spiel believes that “only by cross training with multiple specialties can any one physician bridge the necessary knowledge required to effectively treat the complexities of the human condition.”

The American Board of Interventional Pain Physicians examines and certifies appropriate applicants as diplomats in interventional pain procedures. Dr. Spiel was the first among the few radiologist and interventional pain specialists on the list to be certified by this prestigious board.
  • Endoscopic Laser Discectomy
  • Selective Nerve Root Blocks
  • Transforaminal Epidurals
  • Facet Joint Blocks
  • Caudal Epidurals
  • Sacroiliac Joint Injections
  • Interlaminar Epidurals
  • Radiofrequency Ablation
  • Discography
  • Pulsed Radiofrequency
  • Medial Branch Block
  • Dorsal Column Stimulator Trials
  • Intradiscal Injections
  • Joint Injections
  • Sympathetic Blocks
  • Vertebroplasty
  • Optimesh
  • Reflex Sympathetic Dystrophy
  • Complex Regional Pain Syndrome
  • Diplomate American Board of Radiology
  • Diplomate American Board of Interventional Pain Physicians
  • Fellow of Interventional Pain Practice
  • Diplomate of the Royal College of Physicians and Surgeons of America
  • Diplomate American Academy of Pain Management
  • Member International Spinal Intervention Society
  • Member American Society of Interventional Pain Physicians
  • Member World Institute of Pain
  • Member American Academy of Pain Medicine
  • Endoscopic Laser Discectomy
  • Biologic Vertebral Augmentation utilizing Optimesh
  • Accredited instructor for CME Category 1 courses for Maricopa
  • Integrated Health System
  • Spinal Cord Stimulation- Trials/Permanent
  • Cervical/Thoracic/Lumbar Interventional Injection Techniques
  • Radiofrequency Lesioning
  • Discography and Intradiscal Therapies
  • Epiduroscopy
  • Endovenous Laser Treatment
  • Coblation Nucleoplasty
  • Basic and Advanced Cardiac Life Support
  • International Spinal Intervention Society
  • American Institute of Ultrasound in Medicine
  • American Society of Interventional Pain Physicians
  • American College of Sports Medicine
  • New Jersey Society of Interventional Pain Physicians
  • American Academy of Pain Management
  • American Academy of Pain Medicine
  • American Medical Association: Physician’s Recognition Award forContinuing Medical Education
  • Executive Board Member: American Board of Interventional Pain Physicians
  • National Course Director, Interventional Pain Institute
  • Editorial Board, Pain Physician Journal


Paul Knoepfler, Ph.D.
633A Shriners Hospital
Sacramento Campus

The Knoepfler Lab is interested in epigenetics in stem cells and cancer. We use cutting edge molecular, cellular and developmental biology methods as well as genomic and gene editing technologies to answer key open questions in these areas of research. We are particularly interested now in the roles of three factors normally in stem cells and during tumorigenesis: histone variant H3.3, the MYC family, and DPPA4. How do these factors link the epigenome to cellular behaviors and tissue growth?

Our big picture goal is to impact human health through novel discoveries that catalyze new treatments for cancer and advances in translational stem cell biology.
For more information please visit the Knoepfler lab homepage at: http://www.chromatin.com/
Genomic functions of developmental pluripotency associated factor 4 (Dppa4) in pluripotent stem cells and cancer. Klein RH, Tung PY, Somanath P, Fehling HJ, Knoepfler PSStem Cell Res. 2018 Jul 19;31:83-94. doi: 10.1016/j.scr.2018.07.009. [Epub ahead of print]
Too Much Carrot and Not Enough Stick in New Stem Cell Oversight Trends. Knoepfler PSCell Stem Cell. 2018 Jul 5;23(1):18-20. doi: 10.1016/j.stem.2018.06.004. Epub 2018 Jun 21.
The FDA and the US direct-to-consumer marketplace for stem cell interventions: a temporal analysisKnoepfler PS, Turner LG., Regen Med. 2018 Jan 12. doi: 10.2217/rme-2017-0115. [Epub ahead of print]
ERBB3-Binding Protein (EBP1) is a novel DPPA4 cofactor in human pluripotent cells. Somanath P, Bush KM, Knoepfler PS.Stem Cells. 2018 Jan 12. doi: 10.1002/stem.2776. [Epub ahead of print]

Priyanka Somanath, Rachel Herndon Klein, Paul S. KnoepflerCRISPR-mediated HDAC2 disruption identifies two distinct classes of target genes in human cellsPLOS. 2017 October 5,  https://doi.org/10.1371/journal.pone.0185627

Martinez-Cerdeno V, Barrilleaux B, McDonough A, Ariza J, Yuen B, Somanath P, Le C, Steward C, Horton K, Knoepfler PBehavior of xeno-transplanted undifferentiated human induced pluripotent stem cells is impacted by microenvironment without evidence of tumorsStem Cells Dev. 2017 Jul 10. doi: 10.1089/scd.2017.0059.
To CRISPR and beyond: the evolution of genome editing in stem cells. Chen KY, Knoepfler PS.Regen Med. 2016 Dec;11(8):801-816. PMID: 27905217

Turner L, Knoepfler PSelling Stem Cells in the USA: Assessing the Direct-to-Consumer IndustryCell Stem Cell. 2016 Jun 29. pii: S1934-5909(16)30157-6. doi: 10.1016/j.stem. 2016.06.007. [Epub ahead of print] PubMed PMID: 27374789.
Review of Stem Cell Dialogues by Sheldon Krimsky(1)Knoepfler PSAm J Bioeth. 2016 Apr;16(4):W12-3. doi: 10.1080/15265161.2016.1145756.
When patients reach out, scientists should reach back carefullyKnoepfler PSNat Med. 2016 Mar 3;22(3):230. doi: 10.1038/nm0316-230.
Reviewing post-publication peer reviewKnoepfler P.Trends Genet. 2015 May;31(5):221-3. doi: 10.1016/j.tig.2015.03.006. Epub 2015 Apr 4.
From bench to FDA to bedside: US regulatory trends for new stem cell therapiesKnoepfler PS.Adv Drug Deliv Rev. 2015 Mar;82-83C:192-196. doi: 10.1016/j.addr.2014.12.001. Epub 2014 Dec 7. Review.
Histone H3.3 regulates dynamic chromatin states during spermatogenesis. Yuen BT, Bush KM, Barrilleaux BL, Cotterman R, Knoepfler PSDevelopment. 2014 Aug 19. pii: dev.106450. [Epub ahead of print]
Miz-1 Activates Gene Expression via a Novel Consensus DNA Binding Motif.. Barrilleaux BL, Burow D, Lockwood SH, Yu A, Segal DJ, Knoepfler PSPLoS One. 2014 Jul 1;9(7):e101151. doi: 10.1371/journal.pone.0101151. eCollection 2014.
Epigenetic mechanisms of tumorigenicity manifesting in stem cells. Tung PY, Knoepfler PSOncogene. Jun 16;0. doi: 10.1038/onc.2014.172. [Epub ahead of print]
Histone H3.3 Mutations: A Variant Path to Cancer. BTK Yuen, PS KnoepflerCancer cell 2013 24 (5), 567-574.
Identification of DPPA4 and DPPA2 as a Novel Family of Pluripotency‐Related Oncogenes. PY Tung, NV Varlakhanova, PS KnoepflerSTEM CELLS 2013 31 (11), 2330-2342.
Key Action Items for the Stem Cell Field: Looking Ahead to 2014PS KnoepflerStem cells and development. 2013
Scientists: you really need to get out of the lab moreKnoepfler PSNat Med. 2013 Sep 6;19(9):1086. doi: 10.1038/nm0913-1086.
Chromatin immunoprecipitation assays for myc and N-myc. Barrilleaux BL, Cotterman R, Knoepfler PSThe Myc Gene, 2013 117-133.
Automation of Library Preparation for High-resolution ChIP-seq Profiling. IM Henry, R Cotterman, P Knoepfler, L Comai, RW Kim, H O’Geen. Journal of Biomolecular Techniques. 2013. JBT 24 (Suppl), S45.

Laskowski AI and Knoepfler PSMyc binds the pluripotency factor Utf1 through the basic-helix-loop-helix leucine zipper domainBiochemical and biophysical research communications.2013
Endogenous mammalian histone H3.3 exhibits chromatin-related functions during development. Bush KM, Yuen BT, Barrilleaux BL, Riggs JW, O Geen H, Cotterman R, Knoepfler PSEpigenetics Chromatin. 2013 Apr 9;6(1):7.

Riggs JW, 
Barrilleaux B, 
Varlakhanova N, 
Bush K, 
Chan V, 
and Knoepfler PSInduced pluripotency and oncogenic transformation are related processesStem Cells & Dev. 2013 Jan 1;22(1):37-50.

Meissen JK, Yuen BTK, Kind T, Riggs JW, Barupala DK, KnoepflerPK*, and Fiehn O*. Induced pluripotent stem cells show metabolomic differences to embryonic stem cells in polyunsaturated phosphatidylcholines and primary metabolismPLoS One 2012;7(10):e46770. *co-corresponding authors.

Laskowski AI and KnoepflerPSUtf1: Goldilocks for ESC bivalencyCell Stem Cell 2012; Dec 7;11(6):732-4.

Martínez-Cerdeño V, Lemen JM, Chan V, Wey A, Lin W, Dent SR, and Knoepfler PSN-Myc and GCN5 regulate significantly overlapping transcriptional programs in neural stem cellsPLoS One. 2012;7(6):e39456.
Knoepfler PSKey anticipated regulatory issues for clinical use of human induced pluripotent stem cellsRegenerative Medicine. 2012; Sep;7(5):713-20.

Dominguez-Frutos E., Lopez Hernandez I., Vendrell V., Gallozzi M., Gutsche K., Sharpe J., Knoepfler PS, Eisenman RN, Trumpp A., and Schimmang T. N-myc controls proliferation, morphogenesis and patterning of the inner earJ. NeuroSci. 2011 31(19):7178-89.

Barrilleaux B. and Knoepfler PSInducing iPS cells to escape the dish. Cell Stem Cell. 2011 Aug 5;9(2):103-11.

Varlakhanova NV, Cotterman RF, Bradnam K, Korf I, and Knoepfler PSMyc and Miz-1 have coordinate genomic functions including targeting Hox genes in human embryonic stem cellsEpigenetics & Chromatin. 2011 Nov 4;4(1):20.
Knoepfler PSMy Year As A Stem Cell BloggerNature. 2011 Jul 27;475(7357):425.

Barrilleaux B. and Knoepfler PSOverexpression of oncogenes in human cells using ecotropic lentivirus for enhanced biosafetyJ Vis Exp. 2011 Jul 24;(53). pii: 2822

Domínguez-Frutos E, López-Hernández I, Vendrell V, Neves J, Gallozzi M, Gutsche K, Quintana L, Sharpe J, Knoepfler PS, Eisenman RN, Trumpp A, Giráldez F, Schimmang T. N-myc Controls Proliferation, Morphogenesis, and Patterning of the Inner EarJ. Neurosci 2011 31(19): 7178-7189.

  • Leukemia & Lymphoma Society Special Fellowship, 2002-2005
  • Howard Temin Award, NCI, 2005-2010
  • Brain Tumor Society Award: 2007-8 Steven C. Higgins Leadership Chair of Research
  • March of Dimes Basil O’Conner Starter Scholar Award, 2008-9.
  • CIRM New Faculty Award: 2009-2014




Exosome (vesicle)

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Exosomes are extracellular vesicles (EVs) that are produced in the endosomal compartment of most eukaryotic cells.[1][2][3] The multivesicular body (MVB) is an endosome defined by intraluminal vesicles (ILVs) that bud inward into the endosomal lumen. If the MVB fuses with the cell surface (the plasma membrane), these ILVs are released as exosomes. In multicellular organisms, exosomes and other EVs are present in tissues and can also be found in biological fluids including bloodurine, and cerebrospinal fluid. They are also released in vitro by cultured cells into their growth medium.[4][5][6] Since the size of exosomes is limited by that of the parent MVB, exosomes are generally thought to be smaller than most other EVs, from about 30 to several hundred nanometres (nm) in diameter: around the same size as many lipoproteins but much smaller than cells.[4] Compared with EVs in general, it is unclear whether exosomes have unique characteristics or functions or can be separated or distinguished effectively from other EVs.[1] EVs including exosomes carry markers of cells of origin and have specialized functions in physiological processes, from coagulation and intercellular signalling to waste management.[4] Consequently, there is a growing interest in clinical applications of EVs as biomarkers and therapies alike,[7] prompting establishment of an International Society for Extracellular Vesicles (ISEV) and a scientific journal devoted to EVs, the Journal of Extracellular Vesicles.


First discovered in the maturing mammalian reticulocyte (immature red blood cell),[8] exosomes were shown to participate in selective removal of many plasma membrane proteins[9] as the reticulocyte becomes a mature red blood cell (erythrocyte). In the reticulocyte, as in most mammalian cells, portions of the plasma membrane are regularly internalized as endosomes, with 50 to 180% of the plasma membrane being recycled every hour.[10] In turn, parts of the membranes of some endosomes are subsequently internalized as smaller vesicles. Such endosomes are called multivesicular bodies because of their appearance, with many small vesicles, (ILVs or “intralumenal endosomal vesicles”), inside the larger body. The ILVs become exosomes if the MVB merges with the cell membrane, releasing the internal vesicles into the extracellular space.[11]

Exosomes contain various molecular constituents of their cell of origin, including proteins and RNA. Although the exosomal protein composition varies with the cell and tissue of origin, most exosomes contain an evolutionarily-conserved common set of protein molecules. The protein content of a single exosome, given certain assumptions of protein size and configuration, and packing parameters, can be about 20,000 molecules.[12] The cargo of mRNA and miRNA in exosomes was first discovered at the University of Gothenburg in Sweden.[13] In that study, the differences in cellular and exosomal mRNA and miRNA content was described, as well as the functionality of the exosomal mRNA cargo. Exosomes have also been shown to carry double-stranded DNA.[14]

Exosomes can transfer molecules from one cell to another via membrane vesicle trafficking, thereby influencing the immune system, such as dendritic cells and B cells, and may play a functional role in mediating adaptive immune responses to pathogens and tumors.[15][16] Therefore, scientists that are actively researching the role that exosomes may play in cell-to-cell signaling, often hypothesize that delivery of their cargo RNA molecules can explain biological effects. For example, mRNA in exosomes has been suggested to affect protein production in the recipient cell.[13][17][18] However, another study has suggested that miRNAs in exosomes secreted by mesenchymal stem cells (MSC) are predominantly pre- and not mature miRNAs.[19] Because the authors of this study did not find RNA-induced silencing complex-associated proteins in these exosomes, they suggested that only the pre-miRNAs but not the mature miRNAs in MSC exosomes have the potential to be biologically active in the recipient cells. Multiple mechanisms have been reported to be involved in loading miRNAs into exosomes, including specific motifs in the miRNA sequences, interactions with lncRNAs localized to the exosomes, interactions with RBPs, and post-translational modifications of Ago[20].

Conversely, exosome production and content may be influenced by molecular signals received by the cell of origin. As evidence for this hypothesis, tumor cells exposed to hypoxia secrete exosomes with enhanced angiogenic and metastatic potential, suggesting that tumor cells adapt to a hypoxic microenvironment by secreting exosomes to stimulate angiogenesis or facilitate metastasis to more favorable environment.[21]


Evolving consensus in the field is that the term “exosome” should be strictly applied to an EV of endosomal origin. Since it can be difficult to prove such an origin after an EV has left the cell, variations on the term “extracellular vesicle” are often appropriate instead.[1][22]


Exosomes from red blood cells contain the transferrin receptor which is absent in mature erythrocytes. Dendritic cell-derived exosomes express MHC IMHC II, and costimulatory molecules and have been proven to be able to induce and enhance antigen-specific T cell responses in vivo. In addition, the first exosome-based cancer vaccination platforms are being explored in early clinical trials.[23] Exosomes can also be released into urine by the kidneys, and their detection might serve as a diagnostic tool.[24][25][26] Urinary exosomes may be useful as treatment response markers in prostate cancer.[27][28] Exosomes secreted from tumour cells can deliver signals to surrounding cells and have been shown to regulate myofibroblast differentiation.[29] In melanoma, tumor-derived vesicles can enter lymphatics and interact with subcapsular sinus macrophages and B cells in lymph nodes.[30] A recent investigation showed that exosome release positively correlates with the invasiveness of ovarian cancer.[31] Exosomes released from tumors into the blood may also have diagnostic potential. Exosomes are remarkably stable in bodily fluids strengthening their utility as reservoirs for disease biomarkers.[32][33] Patient blood samples stored in biorepositories can be used for biomarker analysis as colorectal cancer cell-derived exosomes spiked into blood plasma could be recovered after 90 days of storage at various temperatures.[34]

In malignancies such as cancer, the regulatory circuit which guards exosome homeostasis is co-opted to promote cancer cell survival and metastasis.[35][18]

Urinary exosomes have also proven to be useful in the detection of many pathologies, such as genitourinary cancers and mineralocorticoid hypertension, through their protein and miRNA cargo.”[36] [37]

With neurodegenerative disorders, exosomes appear to play a role in the spread of alpha-synuclein, and are being actively investigated as a tool to both monitor disease progression as well as a potential vehicle for delivery of drug and stem cell based therapy.[38]

An online open access database containing genomic information for exosome content has been developed to catalyze research development within the field.[38]

Exosomes and intercellular communication[edit]

Scientists are actively researching the role that exosomes may play in cell-to-cell signaling, hypothesizing that because exosomes can merge with and release their contents into cells that are distant from their cell of origin (see membrane vesicle trafficking), they may influence processes in the recipient cell [39]. For example, RNA that is shuttled from one cell to another, known as “exosomal shuttle RNA,” could potentially affect protein production in the recipient cell.[17][40] By transferring molecules from one cell to another, exosomes from certain cells of the immune system, such as dendritic cells and B cells, may play a functional role in mediating adaptive immune responses to pathogens and tumors.[15][30]

Conversely, exosome production and content may be influenced by molecular signals received by the cell of origin. As evidence for this hypothesis, tumor cells exposed to hypoxia secrete exosomes with enhanced angiogenic and metastatic potential, suggesting that tumor cells adapt to a hypoxic microenvironment by secreting exosomes to stimulate angiogenesis or facilitate metastasis to more favorable environment.[21] It has recently been shown that exosomal protein content may change during the progression of chronic lymphocytic leukemia.[41]

A study hypothesized that intercellular communication of tumor exosomes could mediate further regions of metastasis for cancer. Hypothetically, exosomes can plant tumor information, such as tainted RNA, into new cells to prepare for cancer to travel to that organ for metastasis. The study found that tumor exosomal communication has the ability to mediate metastasis to different organs. Furthermore, even when tumor cells have a disadvantage for replicating, the information planted at these new regions, organs, can aid in the expansion of organ specific metastasis.[42]

Exosomes carry cargo, which can augment innate immune responses. For example, exosomes derived from Salmonella enterica-infected macrophages but not exosomes from uninfected cells stimulate naive macrophages and dendritic cells to secrete pro-inflammatory cytokines such as TNF-α, RANTES, IL-1ra, MIP-2, CXCL1, MCP-1, sICAM-1, GM-CSF, and G-CSF. Proinflammatory effects of exosomes are partially attributed to lipopolysaccharide, which is encapsulated within exosomes[43].


The isolation and detection of exosomes has proven to be complicated.[4][44] Due to the complexity of body fluids, physical separation of exosomes from cells and similar-sized particles is challenging. Isolation of exosomes using differential ultracentrifugation results in co-isolation of protein and other contaminants and incomplete separation of vesicles from lipoproteins. Combining ultracentrifugation with micro-filtration or a gradient can improve purity.[45][46] Single step isolation of extracellular vesicles by size-exclusion chromatography has been demonstrated to provide greater efficiency for recovering intact vesicles over centrifugation,[47] although a size-based technique alone will not be able to distinguish exosomes from other vesicle types. To isolate a pure population of exosomes a combination of techniques is necessary, based on both physical (e.g. size, density) and biochemical parameters (e.g. presence/absence of certain proteins involved in their biogenesis).

Often, functional as well as antigenic assays are applied to derive useful information from multiple exosomes. Well-known examples of assays to detect proteins in total populations of exosomes are mass spectrometry and Western blot. However, a limitation of these methods is that contaminants may be present that affect the information obtained from such assays. Preferably, information is derived from single exosomes. Relevant properties of exosomes to detect include size, density, morphology, composition, and zeta potential.[48]


Since the diameter of exosomes is typically below 100 nm and because they have a low refractive index, exosomes are below the detection range of many currently used techniques. A number of miniaturized systems, exploiting nanotechnology and microfluidics, have been developed to expedite exosome analyses. These new systems include a microNMR device,[49] a nanoplasmonic chip,[50] and an magneto-electrochemical sensor[51] for protein profiling; and an integrated fluidic cartridge for RNA detection.[52] Flow cytometry is an optical method to detect exosomes in suspension. Nevertheless, the applicability of flow cytometry to detect single exosomes is still inadequate due to limited sensitivity and potential measurement artifacts such as swarm detection.[53] Other methods to detect single exosomes are atomic force microscopy,[54] nanoparticle tracking analysis,[55] Raman microspectroscopy,[56] tunable resistive pulse sensing, and transmission electron microscopy.[53]

Bioinformatics analysis[edit]

Exosomes contain RNA, proteins, lipids and metabolites that is reflective of the cell type of origin. As exosomes contain numerous proteins, RNA and lipids, large scale analysis including proteomics and transcriptomics is often performed. Currently, to analyse these data, non-commercial tools such as FunRich[57] can be used to identify over-represented groups of molecules. With the advent of Next generation sequencing technologies, the research on exosomes have been accelerated in not only cancer but various diseases. Recently, bioinformatics based analysis of RNA-Seq data of exosomes extracted from Trypanosoma cruzi has showed the association of these extracellular vesicles with various important gene products that strengthens the probability of finding biomarkers for Chagas disease.[58][59]

Therapeutics and carriers of drugs[edit]

Increasingly, exosomes are being recognized as potential therapeutics as they have the ability to elicit potent cellular responses in vitro and in vivo.[60][61][62] Exosomes mediate regenerative outcomes in injury and disease that recapitulate observed bioactivity of stem cell populations.[63][64] Mesenchymal stem cell exosomes were found to activate several signaling pathways important in wound healing (AktERK, and STAT3) and bone fracture repair.[65][66] They induce the expression of a number of growth factors (hepatocyte growth factor (HGF), insulin-like growth factor-1 (IGF1), nerve growth factor (NGF), and stromal-derived growth factor-1 (SDF1)).[67] Exosomes secreted by human circulating fibrocytes, a population of mesenchymal progenitors involved in normal wound healing via paracrine signaling, exhibited in-vitro proangiogenic properties, activated diabetic dermal fibroblasts, induced the migration and proliferation of diabetic keratinocytes, and accelerated wound closure in diabetic mice in vivo. Important components of the exosomal cargo were heat shock protein-90α, total and activated signal transducer and activator of transcription 3, proangiogenic (miR-126, miR-130a, miR-132) and anti-inflammatory (miR124a, miR-125b) microRNAs, and a microRNA regulating collagen deposition (miR-21).[68] Researchers have also found that exosomes released from oral keratinocytes can accelerate wound healing, even when human exosomes were applied to rat wounds.[69] Exosomes can be considered a promising carrier for effective delivery of small interfering RNA due to their existence in body’s endogenous system and high tolerance.[70][71] Patient-derived exosomes have been employed as a novel cancer immunotherapy in several clinical trials.[72]

Exosomes offer distinct advantages that uniquely position them as highly effective drug carriers. Composed of cellular membranes with multiple adhesive proteins on their surface, exosomes are known to specialize in cell–cell communications and provide an exclusive approach for the delivery of various therapeutic agents to target cells.[73] For example, researchers used exosomes as a vehicle for the delivery of cancer drug paclitaxel. They placed the drug inside exosomes derived from white blood cells, which were then injected into mice with drug-resistant lung cancer. Importantly, incorporation of paclitaxel into exosomes increased cytotoxicity more than 50 times as a result of nearly complete co-localization of airway-delivered exosomes with lung cancer cells.[74]


Ambrosia: closed its doors 2019: https://www.ambrosiaplasma.com/



NEWS | January 19, 2016

Researchers decipher stem cell messages in blood vessel formation

Findings may lead to novel treatment approaches for peripheral artery disease


An international collaboration between UC Davis and Swedish scientists has resulted in the first comprehensive characterization of a recently discovered cell-to-cell communication system used by stem cells. The findings, posted online today in the journal Stem Cells, will help scientists develop new stem cell-based treatment options for peripheral artery disease, a condition which affects about 12 million people in the United States alone.
Johnathon Anderson Johnathon Anderson

Proteins, RNA and other cellular contents were recently discovered to be released from stem cells in millions of small bubbles – called exosomes – and taken up by neighboring cells, influencing their behavior and activity. This latest research has characterized for the first time the proteins contained in exosomes from mesenchymal stem cells by exposing them to both normal physiological conditions and low oxygen conditions that mimic the low blood flow seen in peripheral arterial disease.

“We have provided the first comprehensive snapshot of the communication proteins’ stem cells and their exosomes use to talk to other cells,” said Johnathon Anderson, lead author of the study and a research scientist with the UC Davis Stem Cell Program. “It has enabled us to fundamentally grasp how stem cells use this newly identified communication system to heal damaged tissues that have reduced or cut off blood flow in individuals suffering from peripheral arterial disease.”

The researchers focused on mesenchymal stem cells (MSCs), which are stem cells found in the bone marrow of adults that can renew themselves and repair damaged tissue. According to Anderson, the traditional view of MSCs emphasized their ability to differentiate into different cell types. The more recent view suggests a therapeutic role for these so-called “paramedic cells” that help heal tissue by homing in on damaged tissues and encouraging more cells to help out in the revascularization process.

How MSCs influence other cells has been a source of intense research interest. Until recently, scientists believed that they primarily communicated by simply secreting individual proteins outside of themselves into the intercellular space – the area between cells.

In the last few years, a sophisticated communications system has been discovered involving exosomes – small bubbles that cells use to transport part of themselves to neighboring cells. Cells use the intricate communication system to cope with changes in their surroundings such as low oxygen or stressful conditions.

Anderson worked in collaboration with the Karolisnka Institute’s Janne Lehtio. Their research teams used a specialized mass spectrometry system known as “high-resolution isoelectric focusing coupled liquid chromatography tandem mass spectrometry” (HiRIEF LC-MS/MS), which allowed them to characterize the protein contents of MSCs and MSC-derived exosomes in a comprehensive fashion for the first time. They identified more than 6,000 different proteins in MSCs and nearly 2,000 in exosomes, and through computer algorithms were able to discern the relationships between these proteins and how they likely interact with one another.

Interestingly, out of nearly 2,000 proteins found in exosomes, nearly one-quarter were not detected in the MSCs, indicating that MSCs might be making certain proteins solely for transportation in the communication bubbles.

Investigators also found the paramedic cells (MSCs) reacted very differently when exposed to normal physiological conditions as compared to low-oxygen conditions, which mimic the low blood flow seen in peripheral artery disease.

The research revealed that MSCs exposed to low-oxygen conditions expressed several proteins in higher concentrations, especially those associated with new blood vessel formation – a process called angiogenesis. Most notable was the expression of proteins involved with nuclear factor-kappaB (NFkB) signaling, an important communication pathway that helps new blood vessels form and is a vital process in treating peripheral arterial disease.

“One of the most interesting aspects of these little exosome bubbles is the fact that we can package them with novel types of therapeutics to enhance their tissue-healing capabilities,” said Anderson. “With this fundamental understanding of how the communication system works, it really opens the floodgates for developing new therapeutics for cardiovascular and neurological diseases using MSC exosomes.”

“This research represents a significant step in the field of mesenchymal stem cell communication,” added Jan Nolta, principal investigator of the study and director of UC Davis’ Stem Cell Program and its Institute for Regenerative Cures in Sacramento. “The results help to explain how MSCs and the exosomes that they produce can be such effective mediators of angiogenesis and have the potential to further the development of new treatments for ischemic tissue-related diseases.”

The article in Stem Cells is entitled, “Comprehensive proteomic analysis of mesenchymal stem cell exosomes reveals modulation of angiogenesis via NFkB signaling.”

Other study authors from the UC Davis Stem Cell Program are Calvin Graham, Missy Pham, Charles Bramlett, Elizabeth Montgomery, Renee Bardini, Gerhard Bauer, Kyle Fink, Brian Fury, Zelenia Contreras, Madeline Hoon and Kyle Hendrix. Additional authors include Matt Mellema from the UC Davis Department of Veterinary Medicine; Frederic Chedin from the UC Davis Department of Molecular and Cellular Biology; Billie Hwang and Michael Mulligan from the University of Washington; and  Henrik Johansson, Mattias Versterlund, Janne Lehtiö and Samir EL-Andaloussi from the Karolinska Institutet in Stockholm, Sweden.

The UC Davis research was funded in part by a Transformative Research Award from National Institutes of Health (NIH) Common Fund, for what the agency terms “innovative, unconventional, paradigm-shifting research projects that are inherently risky and untested” (NIH High-Risk, High-Reward Program, R01GM099688, T32-GM008799 and T32-HL086350). Additional research support came from the National Science Foundation (Graduate Research Fellowships Program 2011116000 and Graduate Research Opportunities Worldwide 201111600).
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