This is a very important question. It does appear Adipose (Fat) is a better source of stem cells, in terms of quantity and percentage of key markers of viability, noted in below studies, than Bone Marrow. I did post about this last year in May. http://eyedoc2020.blogspot.com/2017/05/what-is-best-source-of-stem-cells.html
Since then there have been more studies. Though I could find none on the use of Bone Marrow versus Adipose derived Stem Cells for use in regenerating Meibomian Gland cells, there are good studies looking at the benefit of topical (drops only) use of adipose-derived stem cells to heal corneal issues faster than control groups. There are also good articles on adipose derived stem cells to heal lacrimal glands: coming in another post but initial articles are listed below **.
The first Reference below is on the use of Adipose Derived Stem Cells to support the growth of limbal stem cells.
The key finding in general is that adipose derived stem cells are generally safe in animals and humans for certain applications: not all. The complication rate is low for almost all types of injections I could find thus far except injection into the eye vitreous cavity where 3 patients have lost vision from this.
While the animal studies look very promising, there is no 100% guarantee stem cells will work for a particular condition.
Use of human adipose derived stem cells for injection into the orifice of the meibomian gland has not been done yet. It looks promising in general.
We hope to start our adipose derived stem cell injection into the orifice of the meibomian gland hopefully in the next few weeks, pending IRB approval. We appear to have received our IRB approval from Georgetown University's Medical School for the meibomian gland disease issue I have posted about in kids using excessive computers.
Sandra Lora Cremers, MD, FACS
A side note about published studies:
1. I always like to look at which group did the study. Some countries have more rigorous criteria for research than others. Some groups are well known for the quality of their research. Groups from the US are in general more respected.
2. Higher impact factor journals are better usually. I have posted about this also before: Pubmed should have impact journal scores next to titles on the search options.
3. In general: It is best to look at articles that have a control group.
4. All studies need to be reproduced by another independent group before it can be verified.
References:
1. With permission, I have posted the entire article of this paper.
They showed topical stem cell drops healed corneal wounds in mice faster than 100% autologous serum.
Notes:
1. Isolation, preparation, flow cytometry, immunofluorescence, and multilineage differentiation of ADSC have been reported in our previous studies [12,17,18].
12. Beltrami A.P., Cesselli D., Bergamin N., Marcon P., Rigo S., Puppato E., D’Aurizio F., Verardo R., Piazza S., Pignatelli A., et al. Multipotent cells can be generated in vitro from several adult human organs (heart, liver, and bone marrow) Blood. 2007;
PLOS: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0186238
Adipose tissue depots can exist in close association with other organs, where they assume diverse, often non-traditional functions. In stemcell-rich skin, bone marrow, and mammary glands, adipocytes signal to and modulate organ regeneration and remodeling. Skin adipocytes and their progenitors signal to hair follicles, promoting epithelial stem cell quiescence and activation, respectively. Hair follicles signal back to adipocyte progenitors, inducing their expansion and regeneration, as in skin scars. In mammary glands and heart, adipocytes supply lipids to neighboring cells for nutritional and metabolic functions, respectively. Adipose depots adjacent to skeletal structures function to absorb mechanical shock. Adipose tissue near the surface of skin and intestine senses and responds to bacterial invasion, contributing to the body's innate immune barrier. As the recognition of diverse adipose depot functions increases, novel therapeutic approaches centered on tissue-specific adipocytes are likely to emerge for a range of cancers and regenerative, infectious, and autoimmune disorders.
3.
Due to the known disadvantages of autologous bone grafting, tissue engineering approaches have become an attractive method for ridge augmentation in dentistry. To the best of our knowledge, this is the first study conducted to evaluate the potential therapeutic capacity of PRP-assisted hADSCs seeded on HA/TCP granules on regenerative healing response of canine alveolar surgical bone defects. This could offer a great advantage to alternative approaches of bone tissue healing-induced therapies at clinically chair-side procedures.
Cylindrical through-and-through defects were drilled in the mandibular plate of 5 mongrel dogs and filled randomly as following: I- autologous crushed mandibular bone, II- no filling material, III- HA/TCP granules in combination with PRP, and IV- PRP-enriched hADSCs seeded on HA/TCP granules. After the completion of an 8-week period of healing, radiographic, histological and histomorphometrical analysis of osteocyte number, newly-formed vessels and marrow spaces were used for evaluation and comparison of the mentioned groups. Furthermore, the buccal side of mandibular alveolar bone of every individual animal was drilled as normal control samples (n=5).
Our results revealed that hADSCs subcultured on HA/TCP granules in combination with PRP significantly promoted bone tissue regeneration as compared with those defects treated only with PRP and HA/TCP granules (P<0.05).
In conclusion, our results indicated that application of PRP-assisted hADSCs could induce bone tissue regeneration in canine alveolar bone defects and thus, present a helpful alternative in bone tissue regeneration.
The advantages of adipose-derived stem cells (AdSCs) over bone marrow stem cells (BMSCs), such as being available as a medical waste and less discomfort during harvest, have made them a good alternative instead of BMSCs in tissue engineering. AdSCs from buccal fat pad (BFP), as an easily harvestable and accessible source, have gained interest to be used for bone regeneration in the maxillofacial region. Due to scarcity of data regarding comparative analysis of isolated AdSCs from different parts of the body, we aimed to quantitatively compare the proliferation and osteogenic capabilities of AdSCs from different harvesting sites. In this study, AdSCs were isolated from BFP (BFPdSCs), abdomen (abdomen-derived mesenchymal stem cells (AbdSCs)), and hip (hip-derived mesenchymal stem cells (HdSCs)) from one individual and were compared for surface marker expression, morphology, growth rate, and osteogenic differentiation capability. Among them, BFPdSCs demonstrated the highest proliferation rate with the shortest doubling time and also expressed vascular endothelial markers including CD34 and CD146. Moreover, the expression of osteogenic markers were significantly higher in BFPdSCs. The results of this study suggested that BFPdSCs as an encouraging source of mesenchymal stem cells are to be used for bone tissue engineering.
Mesenchymal stem cell (MSC) therapy has emerged as a potential novel method of treating liver fibrosis. To date, bone marrow-derived MSCs (BM-MSCs) and adipose tissue-derived MSCs (AD-MSCs) have not been analyzed with respect to their ability to combat liver fibrosis. The present study aimed to compare the capabilities of BM-MSCs and AD-MSCs in the treatment of liver fibrosis. BM-MSCs and AD-MSCs were taken from male Sprague-Dawley rats and cultured. Hepatic stellate cells (HSCs) were co-cultured with either BM-MSCs or AD-MSCs, and the effects of BM-MSCs or AD-MSCs on the proliferation, activation and apoptosis of HSCs were determined. The secretion of a selected group of cytokines by BM-MSCs and AD-MSCs was measured using enzyme-linked immunosorbent assays. Using a CCl4-induced liver fibrosis animal model, the anti-inflammatory and anti-fibrotic effects of BM-MSCs or AD-MSCs against liver fibrosis in vivo were evaluated. The morphological examination and analysis of specific surface markers confirmed the successful preparation of BM-MSCs and AD-MSCs. Furthermore, the proliferation, activation and apoptosis of HSCs were significantly inhibited by BM-MSCs and AD-MSCs, with statistically greater reductions achieved by AD-MSCs compared with BM-MSCs. Direct comparison of the secretion of selected cytokines by BM-MSCs and AD-MSCs revealed that significantly higher levels of nerve growth factor and transforming growth factor-β1 were secreted in the AD-MSC culture medium, whereas levels of vascular endothelial growth factor and interleukin-10 did not differ significantly between AD-MSCs and BM-MSCs. In vivo studies using a CCl4-induced liver fibrosis model demonstrated that inflammatory activity and fibrosis staging scores were significantly lower in the MSC-treated groups compared with controls. Although AD-MSCs improved anti-inflammatory and anti-fibrotic effects compared with BM-MSCs, these differences were not significant. Thus, the current study demonstrated that BM-MSCs and AD-MSCs are similarly effective at attenuating liver fibrosis by inhibiting the activation and proliferation of HSCs, as well as promoting the apoptosis of HSCs.
In this review, we compared the potential of mesenchymal stem cells derived from bone marrow, adipose tissue and umbilical cord as suitable sources for regeneration of inner ear hair cells and auditory neurons. Our intensive literature search indicates that stem cells in some of adult mammalian tissues, such as bone marrow, can generate new cells under physiological and pathological conditions. Among various types of stem cells, bone marrow-derived mesenchymal stem cells are one of the most promising candidates for cell replacement therapy. Mesenchymal stem cells have been reported to invade the damaged area, contribute to the structural reorganization of the damaged cochlea and improve incomplete hearing recovery. We suggest that bone marrow-derived mesenchymal stem cells would be more beneficial than other mesenchymal stem cells.
Most adipocytes exist in discrete depots throughout the body, notably in well-defined white and brown adipose tissues. However, adipocytes also reside within specialized niches, of which the most abundant is within bone marrow. Whereas bone marrow adipose tissue (BMAT) shares many properties in common with white adipose tissue, the distinct functions of BMAT are reflected by its development, regulation, protein secretion, and lipid composition. In addition to its potential role as a local energy reservoir, BMAT also secretes proteins, including adiponectin, RANK ligand, dipeptidyl peptidase-4, and stem cell factor, which contribute to local marrow niche functions and which may also influence global metabolism. The characteristics of BMAT are also distinct depending on whether marrow adipocytes are contained within yellow or red marrow, as these can be thought of as 'constitutive' and 'regulated', respectively. The rBMAT for instance can be expanded or depleted by myriad factors, including age, nutrition, endocrine status and pharmaceuticals. Herein we review the site specificity, age-related development, metabolic characteristics and regulation of BMAT under various metabolic conditions, including the functional interactions with bone and hematopoietic cells.
Author information
3. Curr Eye Res. 2013 Apr;38(4):451-63. doi: 10.3109/02713683.2012.763100. Epub 2013 Feb 1.
Adipose-derived stem cells (ADSC) are multipotent, safe, non-immunogenic and can differentiate into functional keratocytes in situ. The topical use of ADSC derived from human processed lipoaspirate was investigated for treating injured rat cornea.
A total of 19 rats were used. Six animals initially underwent corneal lesion experiments with 0.5 N NaOH (right eye) and 0.2 N (left). The 0.2 NaOH protocol was then used in 13 rats. All 26 eyes of 13 rats eyes received topical azythromycin bid for 3 d and divided into five treatment groups (n = 5 eyes/group), which included: control, stem cells, serum, stem + serum and adipose (raw human lipoaspirate). The four treatment groups received topical treatment three times daily for 3 d. Stem cells were isolated and harvested from human lipoaspirate. Topical eye drops were prepared daily with 1 × 10(5) cells/treatment. Fluorescein positive defect area and light microscope assessment was performed at 20, 28, 45, 50 and 74 h. Animals were sacrificed at 74 h for histological evaluation. Data were statistically analyzed for differences amongst groups.
The stem cell-treated eyes had significantly smaller epithelial defects at each time point compared to control- and adipose-treated eyes (p < 0.05). This group showed slightly better epithelium healing than the serum and combined group, yet not significantly different. Histology showed that stem cell-treated corneas had complete re-epithelization, with less inflammatory cells and limited fibroblast activation structure compared with the control eyes.
Other Notes:
1.
Autologous fat grafting is commonly used to correct soft-tissue contour deformities. However, results are impaired by a variable and unpredictable resorption rate. Autologous adipose-derived stromal cells in combination with lipoinjection (cell-assisted lipotransfer) seem to favor a long-term persistence of fat grafts, thus fostering the development of devices to be used in the operating room at the point of care, to isolate the stromal vascular fraction (SVF) and produce SVF-enhanced fat grafts with safe and standardized protocols. Focusing on patients undergoing breast reconstruction by lipostructure, we analyzed a standard technique, a modification of the Coleman's procedure, and three different commercially available devices (Lipokit, Cytori, Fastem), in terms of 1) ability to enrich fat grafts in stem cellsand 2) clinical outcome at 6 and 12 months.
To evaluate the ability to enrich stem cells, we compared, for each patient (n=20), the standard lipoaspirate with the respective stem cell-enriched one, analyzing yield, immunophenotype and colony-forming capacity of the SVF cells as well as immunophenotype, clonogenicity and multipotency of the obtained adipose stem cells (ASCs). Regarding the clinical outcome, we compared, by ultrasonography imaging, changes at 6 and 12 months in the subcutaneous thickness of patients treated with stem-cell enriched (n=14) and standard lipoaspirates (n=16).
Both methods relying on the enzymatic isolation of primitive cells led to significant increase in the frequency, in the fat grafts, of SVF cells as well as of clonogenic and multipotent ASCs, while the enrichment was less prominent for the device based on the mechanical isolation of the SVF. From a clinical point of view, patients treated with SVF-enhanced fat grafts demonstrated, at six months, a significant superior gain of thickness of both the central and superior-medial quadrants with respect to patients treated with standard lipotransfer. In the median-median quadrant the effect was still persistent at 12 months, confirming an advantage of lipotransfer technique in enriching improving long-term fat grafts.
This comparative study, based on reproducible biological and clinical parameters and endpoints, showed an advantage of lipotransfer technique in enriching fat grafts in stem cells and in favoring, clinically, long-term fat grafts.
More Notes:
--
Since then there have been more studies. Though I could find none on the use of Bone Marrow versus Adipose derived Stem Cells for use in regenerating Meibomian Gland cells, there are good studies looking at the benefit of topical (drops only) use of adipose-derived stem cells to heal corneal issues faster than control groups. There are also good articles on adipose derived stem cells to heal lacrimal glands: coming in another post but initial articles are listed below **.
The first Reference below is on the use of Adipose Derived Stem Cells to support the growth of limbal stem cells.
The key finding in general is that adipose derived stem cells are generally safe in animals and humans for certain applications: not all. The complication rate is low for almost all types of injections I could find thus far except injection into the eye vitreous cavity where 3 patients have lost vision from this.
While the animal studies look very promising, there is no 100% guarantee stem cells will work for a particular condition.
Use of human adipose derived stem cells for injection into the orifice of the meibomian gland has not been done yet. It looks promising in general.
We hope to start our adipose derived stem cell injection into the orifice of the meibomian gland hopefully in the next few weeks, pending IRB approval. We appear to have received our IRB approval from Georgetown University's Medical School for the meibomian gland disease issue I have posted about in kids using excessive computers.
Sandra Lora Cremers, MD, FACS
A side note about published studies:
1. I always like to look at which group did the study. Some countries have more rigorous criteria for research than others. Some groups are well known for the quality of their research. Groups from the US are in general more respected.
2. Higher impact factor journals are better usually. I have posted about this also before: Pubmed should have impact journal scores next to titles on the search options.
3. In general: It is best to look at articles that have a control group.
4. All studies need to be reproduced by another independent group before it can be verified.
References:
1. With permission, I have posted the entire article of this paper.
They showed topical stem cell drops healed corneal wounds in mice faster than 100% autologous serum.
Notes:
1. Isolation, preparation, flow cytometry, immunofluorescence, and multilineage differentiation of ADSC have been reported in our previous studies [12,17,18].
12. Beltrami A.P., Cesselli D., Bergamin N., Marcon P., Rigo S., Puppato E., D’Aurizio F., Verardo R., Piazza S., Pignatelli A., et al. Multipotent cells can be generated in vitro from several adult human organs (heart, liver, and bone marrow) Blood. 2007;
17. Zeppieri M., Salvetat M.L., Beltrami A.P., Cesselli D., Bergamin N., Russo R., Cavaliere F., Varano G.P., Alcalde I., Merayo J., et al. Human adipose-derived stem cells for the treatment of chemically burned rat cornea: Preliminary results. Curr. Eye Res. 2013;38:451–463. doi: 10.3109/02713683.2012.763100.[PubMed] [Cross Ref]
18. Ferro F., Spelat R., Falini G., Gallelli A., D’Aurizio F., Puppato E., Pandolfi M., Beltrami A.P., Cesselli D., Beltrami C.A., Ambesi-Impiombato F.S. Adipose tissue-derived stem cell in vitro differentiation in a three-dimensional dental bud structure. Am. J. Pathol. 2011;178:2299–2310. doi: 10.1016/j.ajpath.2011.01.055. [PMC free article]
J Clin Med. 2017 Dec; 6(12): 115.
Published online 2017 Dec 5. doi: 10.3390/jcm6120115
PMCID: PMC5742804
PLOS: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0186238
- Published: October 11, 2017
Abstract
The most efficient method to expand limbal stem cells (LSCs) in vitro for clinical transplantation is to culture single LSCs directly on growth-arrested mouse fibroblast 3T3 cells. To reduce possible xenobiotic contamination from 3T3s, primary human adipose-derived stem cells (ASCs) were examined as feeder cells to support the expansion of LSCs in vitro. To optimize the ASC-supported culture, freshly isolated limbal epithelial cells in the form of single cells (SC-ASC) or cell clusters (CC-ASC) were cultured using three different methods: LSCs seeded directly on feeder cells, a 3-dimensional (3D) culture system and a 3D culture system with fibrin (fibrin 3D). The expanded LSCs were examined at the end of a 2-week culture. The standard 3T3 culture served as control. Expansion of SC-ASC showed limited proliferation and exhibited differentiated morphology. CC-ASC generated epithelial cells with undifferentiated morphology in all culture methods, among which CC-ASC in 3D culture supported the highest cell doubling (cells doubled 9.0 times compared to cells doubled 4.9 times in control) while maintained the percentage of putative limbal stem/progenitor cells compared to the control. There were few cell-cell contacts between cultured LSCs and ASCs in 3D CC-ASC. In conclusion, ASCs support the growth of LSCs in the form of cell clusters but not in single cells. 3D CC-ASC could serve as a substitute for the standard 3T3 culture to expand LSCs.
2.
Cell Metab. 2018 Jan 9;27(1):68-83. doi: 10.1016/j.cmet.2017.12.002.
Anatomical, Physiological, and Functional Diversity of Adipose Tissue.
Author information
- 1
- Department of Molecular, Cellular, and Developmental Biology, Yale University, 266 Whitney Avenue, New Haven, CT 06520, USA.
- 2
- Department of Developmental and Cell Biology, University of California, Irvine, 845 Health Sciences Road, Irvine, CA 92697, USA; Sue and Bill Gross Stem Cell Research Center, University of California, Irvine, Irvine, CA 92697, USA; Center for Complex Biological Systems, University of California, Irvine, Irvine, CA 92697, USA.
- 3
- Department of Molecular, Cellular, and Developmental Biology, Yale University, 266 Whitney Avenue, New Haven, CT 06520, USA; Department of Dermatology, Yale School of Medicine, Yale University, New Haven, CT 06520, USA. Electronic address: valerie.horsley@yale.edu.
- 4
- Department of Developmental and Cell Biology, University of California, Irvine, 845 Health Sciences Road, Irvine, CA 92697, USA; Sue and Bill Gross Stem Cell Research Center, University of California, Irvine, Irvine, CA 92697, USA; Center for Complex Biological Systems, University of California, Irvine, Irvine, CA 92697, USA. Electronic address: plikus@uci.edu.
Abstract
3.
Arch Bone Jt Surg. 2017 Nov;5(6):406-418.
Effects of Human Adipose-derived Stem Cells and Platelet-Rich Plasma on Healing Response of Canine Alveolar Surgical Bone Defects.
Shafieian R1, Matin MM1, Rahpeyma A1, Fazel A1, Sedigh HS1, Nabavi AS1, Hassanzadeh H1, Ebrahimzadeh-Bideskan A1.
Author information
- 1
- Department of Anatomy and Cell Biology, School of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.
Abstract
BACKGROUND:
METHODS:
RESULTS:
CONCLUSION:
Stem Cells Int. 2017;2017:2156478. doi: 10.1155/2017/2156478. Epub 2017 Dec 14.
Impact of Tissue Harvesting Sites on the Cellular Behaviors of Adipose-Derived Stem Cells: Implication for Bone Tissue Engineering.
Author information
- 1
- Dental Research Center, Research Institute of Dental Sciences, School of Dentistry, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
- 2
- Department of Tissue Engineering, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
- 3
- Department of Applied Cell Sciences, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
Abstract
Exp Ther Med. 2017 Dec;14(6):5956-5964. doi: 10.3892/etm.2017.5333. Epub 2017 Oct 18.
Comparison of bone marrow-vs. adipose tissue-derived mesenchymal stem cells for attenuating liver fibrosis.
Author information
- 1
- Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital, Wenzhou Medical University, Wenzhou, Zhejiang 325000, P.R. China.
- 2
- Department of Anorectal Surgery, The Sixth Affiliated Hospital of Wenzhou Medical University, Lishui, Zhejiang 325000, P.R. China.
Abstract
Int Tinnitus J. 2017 Dec 1;21(2):122-127. doi: 10.5935/0946-5448.20170023.
Comparison of Three Types of Mesenchymal Stem Cells (Bone Marrow, Adipose Tissue, and Umbilical Cord-Derived) as Potential Sources for Inner Ear Regeneration.
Author information
- 1
- Department of Genetics and Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran.
- 2
- Cellular and Molecular Research Center, Basic Health Sciences Institute, Shahrekord University of Medical Sciences, Shahrekord, Iran.
Abstract
Bone. 2018 Jan 14. pii: S8756-3282(18)30008-5. doi: 10.1016/j.bone.2018.01.008. [Epub ahead of print]
Development, regulation, metabolism and function of bone marrow adipose tissues.
Author information
- 1
- Department of Molecular & Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI, United States.
- 2
- Division of Bone and Mineral Diseases, Department of Medicine, Washington University, Saint Louis, MO, United States.
- 3
- Department of Molecular & Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI, United States. Electronic address: macdouga@umich.edu.
Abstract
Stem Cells Dev. 2012 Sep 20;21(14):2724-52. doi: 10.1089/scd.2011.0722. Epub 2012 May 9.
Same or not the same? Comparison of adipose tissue-derived versus bone marrow-derived mesenchymal stem and stromal cells.
Abstract
Mesenchymal stem/stromal cells (MSCs) comprise a heterogeneous population of cells with multilineage differentiation potential, the ability to modulate oxidative stress, and secrete various cytokines and growth factors that can have immunomodulatory, angiogenic, anti-inflammatory and anti-apoptotic effects. Recent data indicate that these paracrine factors may play a key role in MSC-mediated effects in modulating various acute and chronic pathological conditions. MSCs are found in virtually all organs of the body. Bone marrow-derived MSCs (BM-MSCs) were discovered first, and the bone marrow was considered the main source of MSCs for clinical application. Subsequently, MSCs have been isolated from various other sources with the adipose tissue, serving as one of the alternatives to bone marrow. Adipose tissue-derived MSCs (ASCs) can be more easily isolated; this approach is safer, and also, considerably larger amounts of ASCs can be obtained compared with the bone marrow. ASCs and BM-MSCs share many biological characteristics; however, there are some differences in their immunophenotype, differentiation potential, transcriptome, proteome, and immunomodulatory activity. Some of these differences may represent specific features of BM-MSCs and ASCs, while others are suggestive of the inherent heterogeneity of both BM-MSC and ASC populations. Still other differences may simply be related to different isolation and culture protocols. Most importantly, despite the minor differences between these MSC populations, ASCs seem to be as effective as BM-MSCs in clinical application, and, in some cases, may be better suited than BM-MSCs. In this review, we will examine in detail the ontology, biology, preclinical, and clinical application of BM-MSCs versus ASCs.
J Trauma Acute Care Surg. 2017 Apr 27. doi: 10.1097/TA.0000000000001489. [Epub ahead of print]
Differential inflammatory networks distinguish responses to bone marrow-derived vs. adipose-derived mesenchymal stem cell therapies in vascularized composite allotransplantation.
Author information
From the Department of Surgery (R.Z., Y.V.), Department of Plastic Surgery (S.K.R., V.S.G.), University of Pittsburgh, Pittsburgh, Pennsylvania; Division of Plastic Surgery and Hand Surgery (J.A.P., V.S.G.), University Hospital Zurich, Zurich, Switzerland; and Center for Inflammation and Regenerative Modeling (Y.V.), McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania.
Abstract
BACKGROUND:
Vascularized composite allotransplantation (VCA) is aimed at enabling injured individuals to return to their previous lifestyles. Unfortunately, VCA induces an immune/inflammatory response, which mandates lifelong, systemic immunosuppression, with attendant detrimental effects. Mesenchymal stem cells (MSC) - both adipose-derived (AD-MSC) and bone marrow-derived (BM-MSC) - can reprogram inflammation and have been suggested as an alternative to immunosuppression, but their mechanism of action is as yet not fully elucidated. We sought to gain insights into these mechanisms using a systems biology approach.
METHODS:
PKH26 (red) dye-labeled AD-MSC or BM-MSC were administered intravenously to Lewis rat recipients of mismatched Brown Norway hindlimb transplants. Short course tacrolimus (FK-506) monotherapy was withdrawn at POD 21. Sera were collected at 4, 6, 18 weeks, assayed for 29 inflammatory/immune mediators, and the resultant data were analyzed using Dynamic Network Analysis (DyNA), Dynamic Bayesian Network (DyBN) inference, and Principal Component Analysis (PCA).
RESULTS:
DyNA network complexity decreased with time in AD-MSC rats, but increased in BM-MSC rats. DyBN and PCA suggested mostly different central nodes and principal characteristics, respectively, in AD-MSC vs. BM-MSC rats.
CONCLUSIONS:
AD-MSC and BM-MSC are associated with both overlapping and distinct dynamic networks and principal characteristics of inflammatory/immune mediators in VCA grafts with short course tacrolimus induction therapy. The decreasing inflammatory complexity of dynamic networks in the presence of AD-MSC supports the previously suggested role for regulatory T cells induced by AD-MSC. The finding of some overlapping and some distinct central nodes and principal characteristics suggests the role of key mediators in the response to VCA in general, as well as potentially differential roles for other mediators ascribed to the actions of the different MSC populations. Thus, combined in vivo/in silico strategies may yield novel means of optimizing MSC therapy for VCA
Which is the best method to obtain adipose derived stem cells?
Autologous fat grafting is commonly used to correct soft-tissue contour deformities. However, results are impaired by a variable and unpredictable resorption rate. Autologous adipose-derived stromal cells in combination with lipoinjection (cell-assisted lipotransfer) seem to favor a long-term persistence of fat grafts, thus fostering the development of devices to be used in the operating room at the point of care, to isolate the stromal vascular fraction (SVF) and produce SVF-enhanced fat grafts with safe and standardized protocols. Focusing on patients undergoing breast reconstruction by lipostructure, we analyzed a standard technique, a modification of the Coleman's procedure, and three different commercially available devices (Lipokit, Cytori, Fastem), in terms of 1) ability to enrich fat grafts in stem cellsand 2) clinical outcome at 6 and 12 months.
To evaluate the ability to enrich stem cells, we compared, for each patient (n=20), the standard lipoaspirate with the respective stem cell-enriched one, analyzing yield, immunophenotype and colony-forming capacity of the SVF cells as well as immunophenotype, clonogenicity and multipotency of the obtained adipose stem cells (ASCs). Regarding the clinical outcome, we compared, by ultrasonography imaging, changes at 6 and 12 months in the subcutaneous thickness of patients treated with stem-cell enriched (n=14) and standard lipoaspirates (n=16).
Both methods relying on the enzymatic isolation of primitive cells led to significant increase in the frequency, in the fat grafts, of SVF cells as well as of clonogenic and multipotent ASCs, while the enrichment was less prominent for the device based on the mechanical isolation of the SVF. From a clinical point of view, patients treated with SVF-enhanced fat grafts demonstrated, at six months, a significant superior gain of thickness of both the central and superior-medial quadrants with respect to patients treated with standard lipotransfer. In the median-median quadrant the effect was still persistent at 12 months, confirming an advantage of lipotransfer technique in enriching improving long-term fat grafts.
This comparative study, based on reproducible biological and clinical parameters and endpoints, showed an advantage of lipotransfer technique in enriching fat grafts in stem cells and in favoring, clinically, long-term fat grafts.
Notes to consider:
A. Google search for:
"human adipose derived Stem Cells for lacrimal gland"
Will write another post specifically for this category this week.
derived mesenchymalstem cells for cellular therapy. ... Safety and immunomodulatory effects of allogeneic canine adipose-derived mesenchymal stromal cells transplanted into the region of the lacrimal gland, the gland of the third eyelid and ...
-MSCs) implanted around thelacrimal glands in dogs ( eyes) with KCS, which is refractory to. current available treatments. Schirmer tear test ...
A-C) H&E staining: (A) Normal rat submandibular salivary gland showing mixed glandular tissue. Coagulative necrosis and interstitial edema were observed in the irradiated glands at 24 weeks. (B) The cytoplasm was homogeneously stained ...
hADSCs) can differentiate into salivary gland cells (SGCs) when ...
...
MSC) for the regeneration of lacrimal gland tissue could result in a novel therapy for dry-eye syndrome. To optimize the culture conditions, the purpose of this study was to evaluate the influence of low oxygen on phenotype, differentiation ...
B.
Comparison of Methods for Obtaining and Preparing Adipose Derived Stem Cells from Liposuction.
1. Am J Pathol. 2011 May; 178(5): 2299–2310.
Note:
ADSC were obtained from human adipose tissue aspirates following a protocol optimized by Beltrami’s group for the isolation and in vitro expansion of human multipotent adult stem cells.20
As previously shown for multipotent adults stem cells obtained from human liver, bone marrow, heart and peripheral blood, ADSC expressed the pluripotent state-specific transcription factors Oct-4, Nanog and Sox 2 (Figure 1A–D) and were characterized by a mesenchymal stem cell immunophenotype. When assessed by flow-cytometry, ADSC highly expressed CD90, CD105, CD73, however, were mainly negative for the hematopoietic markers CD34 and CD45 (Figure 1E). Importantly, ADSC displayed multipotency, being able to differentiate into mature cell types of all the three germ layers. Specifically, when exposed to the proper differentiation inducing conditions, ADSC were able to give rise to endodermic (Figures 1F and G), mesodermic (Figures 1H and I) and ectodermic derivatives (Figures 1J and K).
Stem Cell Res Ther. 2015 Jan 5;6:2. doi: 10.1186/scrt536.
Adipose tissue derived stem cells: in vitro and in vivo analysis of a standard and three commercially available cell-assisted lipotransfer techniques.
Domenis R1, Lazzaro L2, Calabrese S3, Mangoni D4, Gallelli A5, Bourkoula E6, Manini I7, Bergamin N8, Toffoletto B9, Beltrami CA10, Beltrami AP11, Cesselli D12, Parodi PC13,14.
Author information
- 1
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. rossana.domenis@uniud.it.
- 2
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. laralazzaro@hotmail.com.
- 3
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. sarah2012@hotmail.com.
- 4
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. damianomng@gmail.com.
- 5
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. byoteck@libero.it.
- 6
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. jenny_bourkoula@hotmail.com.
- 7
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. ivana.manini@tiscali.it.
- 8
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. nataschab@libero.it.
- 9
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. t_barbara2001@yahoo.it.
- 10
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. carloalberto.beltrami@uniud.it.
- 11
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. antonio.beltrami@uniud.it.
- 12
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. daniela.cesselli@uniud.it.
- 13
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. piercamillo.parodi@uniud.it.
- 14
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. piercamillo.parodi@uniud.it.
Abstract
INTRODUCTION:
METHODS:
RESULTS:
CONCLUSIONS:
Notes to consider:
A. Google search for:
"human adipose derived Stem Cells for lacrimal gland"
Will write another post specifically for this category this week.
Scholarly articles for human adipose derived Stem Cells for lacrimal gland | |
… adipose-derived stem cells suppress mixed lymphocyte … - Cui - Cited by 234
… gene delivery into human adipose derived stem cells - Ahn - Cited by 78
… adipose-derived mesenchymal stem cells: an in vivo … - Wood - Cited by 47
|
Search Results
Use of Adipose-Derived Mesenchymal Stem Cells in ... - NCBI - NIH
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4352730/
by AJ Villatoro - 2015 - Cited by 20 - Related articles
Feb 23, 2015 - To our knowledge, this is the first time in literature that implantation of allogeneic Ad-MSCs around lacrimal glands has been found as an effective therapeutic alternative to treat dogs with KCS. These results could reinforce a good effective solution to be extrapolated to future studies inhuman. Go to: ...PubMed Result - NCBI
https://www.ncbi.nlm.nih.gov/pubmed?db=pubmed&cmd=link&linkname...
4: Aghayan HR, Goodarzi P, Arjmand B. GMP-compliant human adipose tissue-Use of Adipose-Derived Mesenchymal Stem Cells in ... - ResearchGate
https://www.researchgate.net/.../274087472_Use_of_Adipose-Derived_Mesenchymal_St...
Dec 20, 2017 - prevalence in humans and dogs. Our aim in this study was to investigate the therapeutic e ects of allogeneic adipose-derived. mesenchymal stromal cells (AdInjury tissue restoration by human adipose tissue-derived stem cells...
https://www.researchgate.net/.../263896466_fig3_Injury-tissue-restoration-by-human-ad...
Injury tissue restoration by human adipose tissue-derived stem cells (hADSCs). (Transdifferentiation of mouse adipose-derived stromal cells into acinar ...
https://www.sciencedirect.com/science/article/pii/S0014482715000919
by J Lee - 2015 - Cited by 9 - Related articles
May 15, 2015 - Huang and colleagues proposed that human amniotic epithelial cells (hAECs) can differentiate into ACs, and the cells are positive for mucins after two weeks of co-culture with ACs [8].Human adipose-derived mesenchymal stem cells (Adult Gland Derived Stem Cells (Gdscs); Potentials, Hurdles and ...
https://www.omicsonline.org/.../adult-gland-derived-stem-cells-gdscs-potentials-hurdle...
A relatively new and particularly unique field of stem cells research due to the rarity of the needed in vitro human specimens for study. The lacrimal gland is made up of acini, ducts, nerves, myoepithelial cells, and plasma cells secreting a tear film interface between the external environment and the ocular surface [95].Fat Grafting: Current Concept, Clinical Application, and ...
https://books.google.com/books?isbn=0323370071
Lee L.Q. Pu - 2015 - Medical
Multipotential human adipose-derived stromal stem cells exhibit a perivascular phenotype in vitro and in vivo. J Cell ... Safety and immunomodulatory effects of allogeneic canine adipose-derived mesenchymal stromal cells transplanted into the region of the lacrimal gland, the gland of the third eyelid and the knee joint.Human adipose tissue-derived stem cells alleviate radiation ...
https://www.semanticscholar.org/.../Human-adipose...derived-stem-cells.../95b4d11c473...
In the present study, we developed a novel approach to regenerate the function of the irradiation‑damaged salivary glands using human adipose tissue‑derived stem cell (hADSC) intraglandular transplantation. ZsGreen‑labeled hADSCs were adoptively transferred into Sprague‑Dawley (SD) rat submandibular glandsThe Influence of Oxygen on the Proliferative Capacity and ... - IOVS
iovs.arvojournals.org/article.aspx?articleid=2411358
by M Roth - 2015 - Cited by 3 - Related articles
Purpose: The application of lacrimal gland–derived mesenchymal stem cells (LG-Adipose-Derived Mesenchymal Stem Cells Reduce Lymphocytic ...
iovs.arvojournals.org/article.aspx?articleid=2565696
by X Li - 2016 - Cited by 1 - Related articles
Primary Sjogren's syndrome (SS) is a chronic autoimmune disease characterized by lymphocytic infiltration of exocrine glands, which leads to functional impairment of the salivary and lacrimal glands.1Whereas the progression of SS has typically been associated with Th1 cell phenotype, new studies have shown that ...Comparison of Methods for Obtaining and Preparing Adipose Derived Stem Cells from Liposuction.
1. Am J Pathol. 2011 May; 178(5): 2299–2310.
PMCID: PMC3081158
Adipose Tissue-Derived Stem Cell in Vitro Differentiation in a Three-Dimensional Dental Bud Structure
Materials and Methods
To isolate adipose tissue–derived stem cells (ASCs), nine raw human abdominal lipoaspirates, obtained with the informed consent of the donors (28 to 35 years old), were washed in PBS solution and then dissociated in Dulbecco's modified Eagle's medium (DMEM; Sigma-Aldrich, St. Louis, MO) containing 400 U/mL of collagenase type 2 (Wortington, Lakewood, NJ) for 15 minutes at 37°C. Samples were centrifuged for 15 minutes at 1,800 × g, then pellets were resuspended and filtered through a 70-μm pore-sized membrane. Filtered cells were plated into 100-mm dishes (2 × 106 cells per dish) in proliferation medium, derived from Gronthos et al,11 composed of F-12 Coon's and Ambesi's modified medium (Gibco-Invitrogen, Carlsbad, CA), Medium-199, and CMRL-1066 media (Sigma) supplemented with 1.25% type 0 human serum, 25 ng/mL of platelet-derived growth factor-bb, 25 ng/mL of epidermal growth factor, 25 ng/mL of insulin-like growth factor 1, 25 ng/mL of fibroblast growth factor-b (FGF-b; all from Immunotools, Friesoythe, Germany), 10−9 mol/L dexamethasone (MP Biomedicals, Solon, OH), 90 μg/L of linoleic acid (Sigma), 25 mg/L of ascorbic acid (Sigma), and 25 μg/mL of gentamicin (Gibco). Colonies developed in primary culture and reached near confluency in approximately 1 week. ASCs were maintained semiconfluent to prevent cell differentiation, and approximately 80% of the medium was replaced every 3 days. Primary human bone marrow mesenchymal stem cells (MSCs) and dental pulp stem cells (DPSCs) were obtained as described by Ferro et al.12 Human bone MSCs were obtained by flushing the femur head content through a 26-gauge needle. Flushed cells were diluted in Hanks' balanced salt solution (Sigma), layered on top of Ficoll (Amersham), and centrifuged. Finally, buffy coat was washed twice with Hanks' balanced salt solution and subjected to immunodepletion using RosetteSep (Stemcells Technologies, Vancouver, British Columbia).13,14 Then 1.5 × 106 of freshly isolated cells, derived from primary culture, were plated in 100-mm dishes (BD Falcon, San Jose, CA). Dental pulps were extracted from human deciduous teeth of 5- to 7-year-old children (with parents' permission) using a syringe needle and were transferred to 35-mm Petri dishes (BD Falcon); human DPSC colonies developed in primary culture and usually reached confluence approximately 2 weeks later; both MSCs and DPSCs were made to proliferate in the same medium used for ASCs.
Human supranumeral teeth buds were isolated following the same methods used for DPSCs; were cultured in DMEM/F-12 (Gibco) containing 20% fetal bovine serum supplemented with 0.18 mg/mL of ascorbic acid (Sigma), 2 mmol/L L-glutamine (Sigma), and 50 U/mL of penicillin/streptomycin (Gibco)8; and were used as amelo-odontoblastic positive control cells. Human primary thyroid cells, used as negative controls in RT-PCR and immunoblot, were cultured according to the method of Curcio et al.15 Human embryonic carcinoma stem cells (Ntera2), used as a positive control for embryonic stem cell markers as suggested by Liedtke et al,16 were cultured in DMEM (Gibco), supplemented with 4.5 g/L of glucose, 4 mmol/L L-glutamine, 10% fetal bovine serum, and 50 U/mL of penicillin/streptomycin (Gibco).17
2.
Am J Pathol. 2011 May; 178(5): 2299–2310.
PMCID: PMC3081158
Adipose Tissue-Derived Stem Cell in Vitro Differentiation in a Three-Dimensional Dental Bud Structure
Federico Ferro,⁎ Renza Spelat,⁎ Giuseppe Falini,† Annarita Gallelli,‡ Federica D'Aurizio,‡ Elisa Puppato,‡ Maura Pandolfi,‡ Antonio Paolo Beltrami,‡ Daniela Cesselli,‡ Carlo Alberto Beltrami,‡ Francesco Saverio Ambesi-Impiombato,⁎and Francesco Curcio⁎⁎Note:
ADSC were obtained from human adipose tissue aspirates following a protocol optimized by Beltrami’s group for the isolation and in vitro expansion of human multipotent adult stem cells.20
As previously shown for multipotent adults stem cells obtained from human liver, bone marrow, heart and peripheral blood, ADSC expressed the pluripotent state-specific transcription factors Oct-4, Nanog and Sox 2 (Figure 1A–D) and were characterized by a mesenchymal stem cell immunophenotype. When assessed by flow-cytometry, ADSC highly expressed CD90, CD105, CD73, however, were mainly negative for the hematopoietic markers CD34 and CD45 (Figure 1E). Importantly, ADSC displayed multipotency, being able to differentiate into mature cell types of all the three germ layers. Specifically, when exposed to the proper differentiation inducing conditions, ADSC were able to give rise to endodermic (Figures 1F and G), mesodermic (Figures 1H and I) and ectodermic derivatives (Figures 1J and K).
3. Curr Eye Res. 2013 Apr;38(4):451-63. doi: 10.3109/02713683.2012.763100. Epub 2013 Feb 1.
Human adipose-derived stem cells for the treatment of chemically burned rat cornea: preliminary results.
Zeppieri M1, Salvetat ML, Beltrami AP, Cesselli D, Bergamin N, Russo R, Cavaliere F, Varano GP, Alcalde I, Merayo J, Brusini P, Beltrami CA, Parodi PC.
Author information
- 1
- Department of Ophthalmology, Azienda Ospedaliero Universitaria Santa Mariadella Misericordia, University of Udine, Udine, Italy. markzeppieri@hotmail.com
Abstract
PURPOSE:
METHODS:
RESULTS:
Isolation and Preparation of Adipose-derived Stem Cells
Human subcutaneous abdominal adipose tissue was obtained from healthy patients (aged 27–62 years) undergoing elective lipoaspiration surgery with informed oral and written consent under a protocol approved by the Institutional Review Board (IRB) of the University of Udine, in accordance with the guidelines of the Tenets of the Declaration of Helsinki. Patients were screened and resulted negative for HIV, hepatitis B and C virus, and syphilis.
ADSC were obtained from the stromal vascular fraction (SVF) obtained from lipoaspirates and cultured as previously described.
Briefly, the SVF was obtained by centrifuging the lipoaspirates at 3000 g for 3 min. The SVF was subsequently dissociated in Jocklik modified Eagle’s medium (JMEM; Sigma-Aldrich, St Louis, MO) containing 400 U/mL of collagenase type 2 (Sigma-Aldrich) for 20 min at 37 °C. The collagenase enzymatic activity was stopped with the addition of 0.1% bovine serum albumin (BSA; Sigma-Aldrich) in JMEM. Samples were centrifuged for 10 min at 600g, then pellets were resuspended and filtered through a 40 µm pore-sized membrane. Filtered cells (2 × 106 cells per dish) were plated into 100 mm human fibronectin-coated (Sigma-Aldrich) dishes in an expansion medium composed as follows: 60% low glucose Dulbecco's Modified Eagle Medium (DMEM; Invitrogen, Carlsbad, CA), 40% MCDB-201(Sigma-Aldrich), 1 mg/mL linoleic acid-BSA, 10−9 mol/L dexamethasone (MP Biomedicals, Solon, OH), 10−4 M ascorbic acid-2 phosphate (Sigma-Aldrich), 1Xinsulin-transferrin-sodium selenite (Sigma-Aldrich), 2% fetal bovine serum (FBS; StemCell Technologies, Vancouver, Canada), 10 ng/mL of human platelet-derived growth factor-bb and10 ng/mL of human epidermal growth factor (both from Peprotech EC, London, UK). Colonies developed in primary culture and reached near confluency in approximately 1 week. Medium was replaced every 3–4 d. Cells were detached with 0.25% trypsin–EDTA (Sigma-Aldrich) and replated at a density of 2 × 103/cm once reached at 70%–80% confluence. Adherent cells obtained after the second subculture, which corresponds to the third passage of cells, were used for the experiment.
Briefly, the SVF was obtained by centrifuging the lipoaspirates at 3000 g for 3 min. The SVF was subsequently dissociated in Jocklik modified Eagle’s medium (JMEM; Sigma-Aldrich, St Louis, MO) containing 400 U/mL of collagenase type 2 (Sigma-Aldrich) for 20 min at 37 °C. The collagenase enzymatic activity was stopped with the addition of 0.1% bovine serum albumin (BSA; Sigma-Aldrich) in JMEM. Samples were centrifuged for 10 min at 600g, then pellets were resuspended and filtered through a 40 µm pore-sized membrane. Filtered cells (2 × 106 cells per dish) were plated into 100 mm human fibronectin-coated (Sigma-Aldrich) dishes in an expansion medium composed as follows: 60% low glucose Dulbecco's Modified Eagle Medium (DMEM; Invitrogen, Carlsbad, CA), 40% MCDB-201(Sigma-Aldrich), 1 mg/mL linoleic acid-BSA, 10−9 mol/L dexamethasone (MP Biomedicals, Solon, OH), 10−4 M ascorbic acid-2 phosphate (Sigma-Aldrich), 1Xinsulin-transferrin-sodium selenite (Sigma-Aldrich), 2% fetal bovine serum (FBS; StemCell Technologies, Vancouver, Canada), 10 ng/mL of human platelet-derived growth factor-bb and10 ng/mL of human epidermal growth factor (both from Peprotech EC, London, UK). Colonies developed in primary culture and reached near confluency in approximately 1 week. Medium was replaced every 3–4 d. Cells were detached with 0.25% trypsin–EDTA (Sigma-Aldrich) and replated at a density of 2 × 103/cm once reached at 70%–80% confluence. Adherent cells obtained after the second subculture, which corresponds to the third passage of cells, were used for the experiment.
Cells were isolated by the use of the selective medium. Stemness of these cells was demonstrated in vitro in accordance to our previous study on the basis of: mesenchymal stem cell-like surface immunophenotype; expression of Oct-4, Nanog and Sox2 proteins and multipotency, shown by the ability to differentiate into derivatives of all three germ layers.
Flow Cytometry
Proliferating cells were detached with 0.25% trypsin–EDTA (Sigma-Aldrich) and, after a 20 min recovery phase, were incubated with either properly conjugated primary antibodies: CD10, CD13, CD29, CD49a, CD49b, CD49d, CD90, CD73, CD44, CD59, CD45, CD271, CD34, (BD Biosciences, Le Pont-de-Claix, France), CD105, KDR, CD66e (Serotech, Kidlington, UK), CD133 (Miltenyi Biotec, Bergisch Gladbach, Germany), E-cadherin (Santa Cruz Biotechnology, Santa Cruz, CA), ABCG-2 (Millipore, Bioscience Research Reagents (formerly Chemicon), Temecula, CA). Properly conjugated isotype-matched antibodies were used as a negative control. The analysis was performed by CyAn (Dako Glostrup, Denmark).
Immunofluorescence
Cells cultured either in expansion or in differentiation medium were fixed in 4% buffered paraformaldehyde for 20 min at room temperature (R.T.). For intracellular stainings, fixed cells were permeabilized for 8 min at R.T. with 0.1% Triton X-100 (Sigma-Aldrich) before exposing them to primary antibodies. In order to block unspecific binding of the primary antibodies, cells were incubated with 10% donkey serum in PBS for 30 min. Primary antibody incubation was performed over-night at 4 °C using following dilutions: Oct-4 (Abcam, 1:150); Sox-2 (Millipore, Bioscience Research Reagents (formerly Chemicon), 1:150); Nanog (Abcam, 1:150); Cytokeratins 7, 8, 18, 19 (Biogenex, Freemont, CA, 1:20); ß3-tubulin (Abcam, 1:1000); Smooth Muscle Actin (Dako, 1:50), Connexin 43 (Santa Cruz, 1:40); α-Sarcomeric Actin (Sigma, 1:100) and Gata4 (Santa Cruz, 1:100). To detect primary antibodies, A488 and A555 dyes labeled secondary antibodies, diluted 1:800, were employed (Molecular Probe, Invitrogen). 0.1 μg/mL DAPI (Sigma-Aldrich) was used to identify nuclei. Vectashield (Vector, Burlingame, CA) was used as a mounting medium. Epifluorescence images were obtained utilizing a live cell imaging dedicated system consisting of a Leica DMI 6000B microscope connected to a Leica DFC350FX camera (Leica Microsystems, Wetzlar, Germany).
Multilineage Differentiation
Hepatocytic differentiation was induced for growing cells for 2 weeks at high density onto fibronectin-coated dishes in a medium containing 0.5% FBS, 10 ng/mL FGF-4 and 20 ng/mL HGF (both from Peprotech EC). After this period, FGF-4 and HGF were substituted for 20 ng/mL OncostatinM for another 14 d (Peprotech EC). Muscle cell differentiation was achieved by plating 0.5 to 1 × 104/cm2 cells in an expansion medium containing 5% FCS (Sigma-Aldrich), 10 ng/mL bFGF, 10 ng/mL VEGF and 10 ng/mL IGF-1 (all from Peprotech EC), but not EGF. Cells were allowed to become confluent and cultured for up to 4 weeks with medium exchanges for every 4 d. For neurogenic differentiation, cells were plated in DMEM-high glucose (Invitrogen), 10% FBS (Sigma-Aldrich). After 24 h medium was replaced with DMEM-high glucose, 10% FBS containing B27 (Invitrogen), 10 ng/mL EGF and 20 ng/mL bFGF (both from Peprotech EC). After 5 d, cells were washed and incubated with DMEM containing 5 g/mL insulin, 200 µM indomethacin and 0.5 mM IBMX (all from Sigma-Aldrich) in the absence of FBS for 5–10 d. At the end of every treatment, cells were fixed with 4% buffered paraformaldehyde.
Experiment for Detecting the Presence of Stem Cells
The preparation of ADSC for treatment was performed in a similar manner to the previous experiments. The cells were allowed to grow until 80% confluence and detached with the trypsin solution, centrifuged and pelleted. The supernatant was discarded and resuspended in a labeling solution containing 25 µm of CFSE in sterile PBS (0.1 M pH 7.4). The cells remained in incubation in this medium for 15 min, then incubated for an additional 30 min with a fresh medium. Cells were washed and centrifuged, then resuspended in HBSS to a final concentration of 5.0 × 106 cells/mL. A small fraction of the cells were plated in a culture dish to demonstrate the viability of the cells after the labeling and the effectiveness of the fluorescent staining.
After 12 h of treatment and one night of rest (24 h after injury), the rat was scarified by an overdose of sodium pentobarbital. Both eyes were enucleated and fixed by immersion in Somogyi's fixative (4% (w/v) paraformaldehyde and 15% (v/v) saturated picric acid solution in phosphate buffer 0.1 M).
The eyeglobes were then cryopreserved in 30% sucrose solution for 2 h and rapidly frozen in liquid nitrogen. We then obtained 5 µm sections on a cryostat microtome. The sections were counterstained with DAPI to label the nuclei and mounted with fluorescent mounting medium (DAKO).
Other Notes:
1.
Stem Cell Res Ther. 2015 Jan 5;6:2. doi: 10.1186/scrt536.
Adipose tissue derived stem cells: in vitro and in vivo analysis of a standard and three commercially available cell-assisted lipotransfer techniques.
Domenis R1, Lazzaro L2, Calabrese S3, Mangoni D4, Gallelli A5, Bourkoula E6, Manini I7, Bergamin N8, Toffoletto B9, Beltrami CA10, Beltrami AP11, Cesselli D12, Parodi PC13,14.
Author information
- 1
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. rossana.domenis@uniud.it.
- 2
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. laralazzaro@hotmail.com.
- 3
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. sarah2012@hotmail.com.
- 4
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. damianomng@gmail.com.
- 5
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. byoteck@libero.it.
- 6
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. jenny_bourkoula@hotmail.com.
- 7
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. ivana.manini@tiscali.it.
- 8
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. nataschab@libero.it.
- 9
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. t_barbara2001@yahoo.it.
- 10
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. carloalberto.beltrami@uniud.it.
- 11
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. antonio.beltrami@uniud.it.
- 12
- Department of Medical and Biological Sciences, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. daniela.cesselli@uniud.it.
- 13
- Clinic of Plastic and Reconstructive Surgery of Udine, University of Udine, P.le Kolbe 4, 33100, Udine, Italy. piercamillo.parodi@uniud.it.
- 14
- Azienda Ospedaliero-Universitaria of Udine, P.le S. Maria della Misericordia 15, 33100, Udine, Italy. piercamillo.parodi@uniud.it.
Abstract
INTRODUCTION:
METHODS:
RESULTS:
CONCLUSIONS:
4.5. Angiogenesis Regulation
Angiogenesis is defined as the process by which new vasculature sprouts from pre-existing blood
vessels. Normal angiogenesis is important during wound healing process. Various studies have
demonstrated the effect of MSC secretome on key steps in angiogenesis. For example, different MSC
populations (e.g., adipose, amniotic, bone marrow (BM) and Wharton jelly umbilical vein) induce
proliferation and migration of endothelial cells promoting tube formation, as well as prevent
endothelial cell apoptosis in vitro [109].
From:
International Journal o f
Molecular Sciences
Review
Mesenchymal Stem Cell Secretome: Toward Cell-Free
Therapeutic Strategies in Regenerative Medicine
Francisco J. Vizoso 1,*, Noemi Eiro 1, Sandra Cid 1, Jose Schneider 2 and
Roman Perez-Fernandez 3,*
1 Research Unit, Fundación Hospital de Jove, Avda. Eduardo Castro, 161, 33290 Gijón, Spain;
noemi.eiro@gmail.com (N.E.); investigacion@hospitaldejove.com (S.C.)
2 Department of Obstetrics & Gynecology, C/Ramon y Cajal 7, University of Valladolid, 47005 Valladolid,
Spain; jose.schneider@urjc.es
3 Department of Physiology-Center for Research in Molecular Medicine and Chronic Diseases (CIMUS),
University of Santiago de Compostela, 15706 Santiago de Compostela, Spain
* Correspondence: franvizoso@gmail.com (F.J.V.); roman.perez.fernandez@usc.es (R.P.-F.);
Tel.: +34-985-320-050 (F.J.V.); +34-881-815-421 (R.P.-F.); Fax: +34-985-315-710 (F.J.V.)
Received: 28 July 2017; Accepted: 22 August 2017; Published: 25 August 2017
Abstract: Earlier research primarily attributed the effects of mesenchymal stem cell (MSC) therapies to
their capacity for local engrafting and differentiating intomultiple tissue types.However, recent studies
have revealed that implanted cells do not survive for long, and that the benefits of MSC therapy
could be due to the vast array of bioactive factors they produce, which play an important role in the
regulation of key biologic processes. Secretome derivatives, such as conditioned media or exosomes,
may present considerable advantages over cells for manufacturing, storage, handling, product shelf
life and their potential as a ready-to-go biologic product. Nevertheless, regulatory requirements for
manufacturing and quality control will be necessary to establish the safety and efficacy profile of
these products. Among MSCs, human uterine cervical stem cells (hUCESCs) may be a good candidate
for obtaining secretome-derived products. hUCESCs are obtained by Pap cervical smear, which is
a less invasive and painful method than those used for obtaining other MSCs (for example, from bone
marrow or adipose tissue). Moreover, due to easy isolation and a high proliferative rate, it is possible
to obtain large amounts of hUCESCs or secretome-derived products for research and clinical use.
Keywords: conditioned media; exosomes; mesenchymal stem cells; adipose-derived stem cells;
bone marrow mesenchymal stem cells; uterine cervical stem cells; hUCESCs
Sandra Lora Cremers, MD, FACS
Johns Hopkins University Medicine, Suburban Hospital
Office: 301-896-0890
*One Central Plaza. 11300 Rockville Pike, Suite 1202. Rockville, MD 20852
*Van Ness Center. 4301 Connecticut Ave., NW, Suite 125. Washington, DC 20008
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Attention: This communication may contain protected health information (PHI) that is legally protected from inappropriate disclosure by the Privacy Standards of the Health Insurance Portability Act (HIPAA) and relevant Maryland Laws. If you are not the intended recipient, please note that any dissemination, distribution or copying of this communication is strictly prohibited. If you have received this message in error, you should notify the sender immediately by telephone or by return e-mail and delete this message from your computer.
Johns Hopkins University Medicine, Suburban Hospital
Office: 301-896-0890
*One Central Plaza. 11300 Rockville Pike, Suite 1202. Rockville, MD 20852
*Van Ness Center. 4301 Connecticut Ave., NW, Suite 125. Washington, DC 20008
www.voeyedr.com
eyedoc2020.blogspot.com
Attention: This communication may contain protected health information (PHI) that is legally protected from inappropriate disclosure by the Privacy Standards of the Health Insurance Portability Act (HIPAA) and relevant Maryland Laws. If you are not the intended recipient, please note that any dissemination, distribution or copying of this communication is strictly prohibited. If you have received this message in error, you should notify the sender immediately by telephone or by return e-mail and delete this message from your computer.
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