TABLE 2.
Studies regarding mesenchymal stromal cell-conditioned medium for treating diabetic wounds in animal models.
| MSC source | Method of tissue extraction | MSC characterization | Preparation of MSC-CM | Model | Groups of treatments and via of administration | Follow-up (days) | Assessment | Main outcome | Other outcomes | |
| Zhang S. et al., 2020 | Human umbilical cords | Full-term delivered infants | Flow cytometry (CD105+, CD73+, CD90+, CD166+, CD54+, CD13+, CD45−, CD34−, CD14−, CD19−, CD117−, HLA-DR−) | MSCs of passages 3–4 at 80% confluence were used. CM was collected | Murine. Diabetic model. Full-thickness excisional skin wounds, 800 mm2, on the back | 150 μl subcutaneously injected around the wounds at six injection sites (25 μl per site) every day for three consecutive days - Control (non-CM) - FB - UC-MSC - UC-MSC-CM (n = 12/group) |
14 | Macroscopic appearance (photography), histology (HE), IHC, qRT-PCR | UC-MSC and UC-MSC-CM treatment accelerated wound healing rate. The wound area of UC-MSC and UC-MSC-CM was ≈15% reduced compared with controls | UC-MSC and UC-MSC-CM increased the percentage of M2 macrophages in the local wounds and the levels of anti-inflammatory cytokines, IL-10, and VEGF and significantly decreased the levels of proinflammatory cytokines, IL-1β, TNF-α, and IL-6 |
| Saheli et al., 2020 | Human bone marrow | − | Flow cytometry (CD73+, CD90+, CD105+, CD34−, CD45−). Osteogenic and adipogenic differentiation | MSCs of passage 4 at 80% were used. CM was then collected | Murine. Diabetic model. Full-thickness excisional skin wounds, 20 mm long, on the chest proximal part intraperitoneal | 50-fold concentrated DMEM or CM twice at 12 and 24 h after wounding intraperitoneally - Control (no treatment) - Placebo - hBM-MSC-CM (n = 6/group) |
15 | Macroscopic appearance, histology (HE, MT), qRT-PCR | MSC-CM accelerated diabetic wound closure (67% wound closure in CM group vs. 33% in placebo group, p < 0.05) | MSC-CM treatment leads to upregulation of EGF and bFGF genes and higher cell viability/proliferation and migration |
| De Gregorio et al., 2020 | Human subcutaneous adipose tissue samples from abdominal region | Liposuction | Flow cytometry (CD29+, CD13+, CD105+, CD73+, CD90+, CD235a−, CD31−, CD45−). Adipogenic and osteogenic differentiation | MSCs of passage 3 at 70% confluence were used. Cells were supplemented with 400 μM DFX (preconditioned MSCs) or with saline (vehicle) as non-preconditioned MSCs. CM was then collected | Murine. Diabetic mice. Full-thickness skin (2.5 mm × 3.5 mm) surgically removed from the dorsal surface of both feet, mimicking a foot ulcer | Intravenous administration of 50 μl of CM every 2 weeks - CM derived from DFX-preconditioned MSCs - CM derived from non-preconditioned MSCs - Vehicle (n = 6/group) |
14 | Macroscopic appearance (photography), histology (HE, MT), qRT-PCR, proteomic analysis | MSC-CM accelerated wound healing. The wound area of MSC-CM was ≈20% reduced compared with controls at day 7 | MSC-CM derived from DFX-preconditioned MSCs had a more potent effect in recovering the skin vasculature |
| Bian et al., 2020 | Human placenta | Cesarean section births | Flow cytometry (CD73+, CD90+, CD105+, CD19−, CD34−, CD45−, HLA-DR−). Osteogenic, adipogenic, and chondrogenic differentiation | MSCs of passages 3–7 were used. CM was collected, and EVs were obtained | Murine. Diabetic model. Full-thickness excisional wounds, 16 mm, on the back | 100 μl was injected around the wounds at 4 sites (25 μl per site) - MSC-EVs - PBS (n = 5/group) |
28 | Photography, histology (HE, MT), IHC | MSC-EVs significantly accelerated wound healing. The narrowest scar widths were observed at day 14 post-wounding (2.41 ± 0.24 mm in MSC-EVs group vs. 3.87 ± 0.60 mm in PBS group) | CXCR4, p21 PCNA, and α-SMA were upregulated in the MSC-EV group |
| Raj et al., 2019 | Human umbilical cord | Umbilical cord dissection | Flow cytometry (CD13+, CD29+, CD44+, CD90+, CD10−, CD14−, CD34−, CD117−) | MSCs were transduced with a lentiviral vector (green fluorescence protein tagged). MSCs of passages 3–4 were used. CM was collected. Wound dressing patches: impregnated with aloe verapolycaprolactone (AV/PCL) nanoscaffolds with hWJSCs or hWJSC−CM were also created | Murine. Diabetic model. Full-thickness excisional wounds, 6 mm, on the back | 100 μl of: - PBS with 1 × 106 MSCs - MSC-CM - UCM - PBS with 1 × 106 MSCs+AV/PCL - MSC-CM+AV/PCL - UCM+ AV/PCL - Untreated group (n = 12/group) |
28 | Macroscopic appearance (photography), histology (HE), IHC, qRT−PCR | Thickness of the epidermis and dermis was significantly greater in both MSCs and MSC-CMs without AV/PCL compared with their controls without AV/PCL | AV/PCL groups showed an earlier re-epithelialization and increases in thickness of dermis and epidermis, cellularity, vasculature, and hair follicle numbers |
| Li T. et al., 2019 | Murine adipose tissue | − | Flow cytometry and oil red O staining (CD29+, CD90, CD45−) | MSCs of passages 3–5 were used. They were cultured on various matrices (tissue culture plates (TCP), pure three-dimensional-printed | Murine. Diabetic murine model. Full-thickness skin defects, 7 mm diameter | 200 μl of each solution was used (150 μl injected subcutaneously around the defect and 50 μl smeared onto the wound bed) - DMEM (control) |
14 | Macroscopic appearance (photography), histology (HE, MT), IF | All groups improved wound healing. The highest wound-healing rate was observed in DOPA-BC-CM group. Remaining wound area at | The newly formed capillary network around the excisional regions was the most intense in the DOPA-BC-CM group. Higher levels of CD31 and a higher amount of |
| bioceramic (BC), and polydopamine-modified BC scaffolds (DOPA-BC), and each CM was collected | - MSC-CMs derived from TCP - MSC-CMs derived from BC - MSC-CMs derived from DOPA-BC (n = 4/group) |
day 14 was 7.1 ± 3.4% in DOPA-BC-CM group, ∼15.2 ± 6.6% in BC-CM, ∼21.2 ± 11.3% in TCP-CM, and ∼31.8 ± 7.2% in DMEM group | collagen deposition were also observed in this group | |||||||
| Deng et al., 2019 | Human adipose tissue | Liposuction | − | SVF gel was prepared, and its CM was collected (Gel-CM). CM from MSCs was also collected (MSC-CM) | Murine. Diabetic model. Full- thickness excisional wound, 20-mm diameter, on the back | 100 μl was administered intradermally every 2 days: - Gel-CM - MSC-CM - PBS (n = 18/group) |
14 | Macroscopic appearance (photography), histology (HE), ELISA | The wound size in all groups was reduced. Wound-healing rate in the Gel-CM-treated group was significantly higher than that in the MSC-CM group (p < 0.05) | Gel-CM-treated rats exhibited complete re-epithelialization of the wound, while MSC-CM did not. Number of capillaries in the Gel-CM-treated group was higher in MSC-CM |
| Dalirfardouei et al., 2019 | Human menstrual blood | Collecting menstrual blood from healthy women | Flow cytometry (CD29+, CD44+, CD90+, CD34−, CD45−, CD117−, HLA−DR-). Adipogenic and osteogenic differentiation | MSCs of passages 4–6 at 70–80% confluence were used. CM was collected, and exosomes were isolated | Murine. Diabetic mice models. Full-thickness excisional wound including the panniculus carnosus, 8 mm, on the back | 100 μl was intradermally injected around the wound - PBS - MSCs - Exos-MSCs (n = 6/group) |
14 | Macroscopic appearance (photography), histology (HE, MT), IHC, qRT-PCR | Increased wound closure was observed in Exo-group compared with the control or MSC group. At day 12, wound closure was 84.34 ± 7.00% in Exo-MSCs, 46.4 ± 8.5% in MSCs, and 43.78 ± 6.95% in controls | Microvessel density was significantly higher in the Exo-group compared with the other two groups. Size of scar tissues significantly decreased in themice treated with MSCs and their exosomes compared with control group. A major reduction in the granulation tissue cellularity was observed in mice treated with exosomes compared with the cell group |
| Irons et al., 2018 | Adipose tissue from pigs’ gluteal regions | Wound incision | − | MSCs at 90% were used. CM was collected | Pigs. Diabetic pigs. Full-thickness skin wounds, 50 mm circular, on the back | -Injection of low-dose MSCs - Injection of high-dose MSCs - Injection of low-dose EC/MSCs - Injection of high-dose EC/MSCs - 2 ml of MSC-CM topically applied every 3 days - 2 ml of EC-CM topically applied every 3 days - 2 ml of serum-free medium topically applied every 3 days (control) (n = 7/group) |
28 | Macroscopic appearance (photography), histology (HE) | Wounds treated with MSCs and MSC-CMs displayed a significant increase in the percentage of wound closure compared with controls (p < 0.05) | Decreases in the acute inflammation scores were observed in wounds treated with MSCs and MSC-CMs compared with controls |
| Chen Z. et al., 2018 | Human umbilical cord | Umbilical cords sections | Electron microscopy (CD63+, CD81+) | MSCs of passages 3–5 at 100% confluence were used. CM was collected, and Exos were obtained. PF-127 hydrogel was mixed with Exos, and PF-127 composite (MSC-Exos/PF-127) was obtained | Murine. Diabetic rat model. Full-thickness skin wounds, 10 mm circular, on the back | Treatment was used topically - 100 μg MSC-Exos dissolved in 100 μl Pluronic F127 hydrogel (24%) - 100 μg MSC-Exos dissolved in 100 μl PBS - 100 μl PF-127 hydrogel (24%) - 100 μl PBS (control) (n = 6/group) |
14 | Macroscopic appearance (photography), histology (HE), IHC, qRT-PCR, IF | Wound area was significantly smaller in the MSC-Exos/PF-127 group than in the other groups. Wounds in the MSC-Exos/PF-127 group were almost completely healed at day 14, while the wound healing rates in the MSC-Exos, PF-127 hydrogel, and control groups were 8.95, 14.52, and 23.09%, respectively | New hair was only evident in the MSC-Exos/PF-127 group. Number of blood vessels was higher in the MSC-Exos/PF-127 and MSC-Exos groups than in the PF-127 hydrogel or control group |
| Chen C. Y. et al., 2018 | Human urine samples | Urine sample collection | Flow cytometry, and electron microscopy (CD29+, CD44+, CD73+, CD90+, CD34−, CD45−). Osteogenic, adipogenic, and chondrogenic differentiation | MSCs of passages 2–6 at 80–90% confluence were used. CM was collected, and Exos were created. Lentivirus shRNAs were transfected | Murine. Diabetic rat model. Full-thickness skin wounds, 6 mm, on upper back | Treatment was subcutaneously injected around the wounds at 4 injection sites (25 μl per site) - 100 μl PBS - 200 μg MSC-Exos in 100 μl PBS - 200 μg MSC-Exos without DMBT1 in 100 μl PBS (n = 8/group) |
12 | Macroscopic appearance (photography), histology (HE, MT), IHC, qRT-PCR, IF | Faster wound closure was observed in MSC-Exos group compared with controls and MSC-Exos without DMBT1 | Higher rate of re-epithelialization, lower level of scar formation, and higher number of newly formed blood vessels were observed in MSC-Exos group compared with controls and MSC-Exos without DMBT1 |
| Bagheri et al., 2018 | Human bone marrow | Aspiration | Flow cytometry (CD73+, CD90+, CD105+, CD34−, CD45−) | MSCs of passage 4 at 80% confluence were used. CM was collected | Murine. Diabetic rat model. Full-thickness skin wounds, 12 mm, on upper back | PBM was administered once daily, 6 days per week. CM was administered at days 0 and 1 intraperitoneally - DMEM vehicle (control) - MSC-CM - PBM - PMB+MSC-CM (n = 18/group) |
15 | Stereological methods, tensiometric examination | MSC-CM and PBM+MSC-CM increased the tensiometric properties compared with DMEM and PBM | MSC-CM, PBM, and PBM+MSC-CM groups showed a significant decrease in the three types of mast cells and in the total number of mast cells compared with controls |
| Amini et al., 2018 | Human bone marrow | Aspiration | Flow cytometry (CD73+, CD90+, CD105+, CD34−, CD45−) | MSCs of passage 4 at 80% confluence were used. CM was collected | Murine. Diabetic rat model. Full-thickness skin wounds, 12 mm, on upper thoracic and lumbar regions | PBM was administered once daily, 6 days per week. CM was administered at days 0 and 1 intraperitoneally - DMEM vehicle (control) - MSC-CM - PBM - PMB+MSC-CM (n = 18/group) |
15 | Stereological methods, tensiometric examination, qRT-PCR | All treated groups significantly enhanced wound healing compared with controls. The extent of healing was significantly greater in the CM+PBM group | Number of fibroblast and epidermal cells, the lengths of blood vessels, and bFGF and SDF-1α expression were significantly higher in the CM+PBM group |
| Pouriran et al., 2016 | Human bone marrow | Aspiration | Flow cytometry (CD105+, CD90+, CD73+, CD34−, CD45−) | MSCs of passage 4 at 80% confluence were used. CM was then collected | Murine. Diabetic rat model. Full-thickness skin wounds, 12 mm, on the thoracic and lumbar regions | PWLLLT was administered once daily, 6 days per week. MSC-CM was administered twice intraperitoneally - Non-treated - MSC-CM - PWLLLT - MSC-CM+PWLLLT (n = 7/group) - Cream containing 1 ml MSC-CM in ratio 10 g cream base - Povidone iodine (control) (n = 6/group) |
15 | Macroscopic appearance (photography), biomechanical examination | PWLLLT and MSC-CM, alone or in combination, improved biomechanical parameters in the wound | PWLLLT was more effective compared with MSC-CM |
| Kusindarta et al., 2016 | Human umbilical cord | − | − | MSCs of passage 4 at 60% confluence were used. CM was then collected | Murine. Diabetic rat model. Full-thickness skin wounds, 7 mm, on the left side of the body | Topical application twice daily | 9 | Macroscopic appearance (photography), histology (HE) | MSC-CM induced faster re-epithelialization than other groups | MSC-CM promoted increasing density of collagen fiber and stimulated hair follicle and muscle regeneration greater than the other groups |
| Ma et al., 2015 | Human hair follicle | Dissection from excess scalp tissue discarded after surgery | Flow cytometry (CD105+, CD29+, CD49b+, CD49d+, CD73+, CD271+, GD2+, CD90−, CD44−, CD34−, CD45−). Adipogenic, osteogenic and chondrogenic differentiation | MSCs of passage 1 at 80–90% confluence were used. CM was then collected | Murine. Diabetic rat model. Full-thickness skin wound, 6 mm, on dorsal surface | 100 ml injected into each wound - DMEM - Normal fibroblast-CM - HF-MSC-CM (n = 3/group) |
24 | Macroscopic appearance (photography), histology (HE) | HF-MSC-CM accelerated wound healing compared with the other groups. The average number of days to complete wound closure in the group administered with HF-MSC-CM was 18.7 days compared with | The epidermal thickness of the HF-MSC-CM-treated wound sites was significantly higher than the other groups |
| 22.3 days in the group treated with fibroblast-CM and 24 days in DMEM group | ||||||||||
| Tam et al., 2014 | Wharton’s jelly from human umbilical cord | Cutting umbilical cords | Flow cytometry (CD13+, CD29+, CD44+, CD90+, CD10−, CD14−, CD34, CD117) | MSCs of passage 3–4 were used. It was constructed wound dressing patch made up of an aloe vera−PCL (AV/PCL) nanoscaffold impregnated with WJ-MSCs or its CM | Murine. Full-thickness skin wounds, 6 mm, on dorsal region | - MSCs+AV/PCL - MSC-CM+AV/PCL - UCM + AV/PCL (n = 12/group) |
28 | Macroscopic appearance (photography), histology (HE and MT), IF, WB, qRT-PCR | MSCs+AV/PCL and MSC-CM+AV/PCL groups showed faster wound closure compared with other groups | MSCs+AV/PCL and MSC-CM+AV/PCL groups showed increased numbers of sebaceous glands and hair follicles and greater cellularity and vasculature compared with other groups |
| Fong et al., 2014 | Wharton’s jelly from human umbilical cord | Full-term delivery | Immunofluorescence (CD10+, CD13+, CD29+, CD44+, CD90+) | MSCs of passages 3–4 at 80% confluence were used. CM was then collected | Murine. Full-thickness excisional skin wound, 6 mm, on dorsum | 100 μl injected intraperitoneally - MSCs - MSC-CM - UCM (n = 12/group) |
28 | Macroscopic appearance (photography), histology (HE), IHC | MSC and MSC-CM healing rates were greater compared with controls | MSCs and MSC-CMs showed greater re-epithelialization, increased vascularity, cellular density, sebaceous gland, and hair follicle numbers compared with controls |
| Kim et al., 2010 | Human umbilical cord | Not specified (CD34+, CD31+, KDR+, Tie2+) | Cells were cultured, and CM was then collected | Murine. Diabetic rats. Full-thickness excisional wounds, 5 mm, on dorso-lateral area | Intradermal injections injected at three different intact dermis site near the wound - MSC - MSC-CM - PBS (n = 6/group) |
12 | Macroscopic appearance (photography), histology (HE) | MSC-CM and MSC promoted wound healing greater than controls | MSC-CM and MSC groups showed greater increases in neovascularization compared with controls. The effect of MSC and MSC-CM improving wound healing was similar |
AT, adipose tissue-derived; bFGF, basic fibroblast growth factor; AF, amniotic fluid; BM, bone marrow; CM, conditioned medium; DFX, deferoxamine; DMEM, Dulbecco’s modified Eagle medium; DP, dental pulp; EC, endothelial-differentiated; ECM, extracellular matrix; EGF, epidermal growth factor; ELISA, enzyme-linked immunosorbent assay; EVs, extracellular vesicles; Exos; exosomes; HE, hematoxylin and eosin; hCB, human cord blood; hESC, human embryonic stem cell; IHC, immunohistochemistry; IF, immunofluorescence; MPF, micro-nano polylactic acid electrospun fiber; MSCs, mesenchymal stromal cells; MT, Masson’s trichrome; PBM, photobiomodulation; PBS, phosphate-buffered saline; qRT-PCR, real-time quantitative polymerase chain reaction; SVF, stromal vascular fraction; UCM, unconditioned medium; WB, Western blotting; WJ, Wharton’s jelly.