Advances in the applications of mesenchymal stem cell-conditioned medium in ocular diseases.

Mesenchymal stem cell-conditioned medium (MSC-CM), also known as secretome, is secreted by MSC and contains a variety of bioactive factors with anti-inflammatory, anti-apoptotic, neuroprotection, and proliferation effects. Increasing evidence proved that MSC-CM plays an important role in various diseases, including skin, bone, muscle, and dental diseases. However, the role of MSC-CM in ocular diseases is not quite clear, Therefore, this article reviewed the composition, biological functions, preparation, and characterization of MSC-CM and summarized current research advances in different sources of MSC-CM in corneal and retinal diseases, including dry eye, corneal epithelial damage, chemical corneal injury, retinitis pigmentosa (RP), anterior ischemic optic neuropathy (AION), diabetic retinopathy (DR), and other retinal degenerative changes. For these diseases, MSC-CM can promote cell proliferation, reduce inflammation and vascular leakage, inhibit retinal cell degeneration and apoptosis, protect corneal and retinal structures, and further improves visual function. Hence, we summarize the production, composition and biological functions of MSC-CM and focus on describing its mechanisms in the treatment of ocular diseases. Furthermore, we look at the unexplored mechanisms and further research directions for MSC-CM based therapy in ocular diseases.

Therapeutic effects of L-arginine, L-carnitine, and mesenchymal stem cell-conditioned medium on endometriosis-induced oocyte poor quality in an experimental mouse model.

AIM: The present study aimed to explore the potential ameliorative effects of L-arginine (LA), L-carnitine (LC), and bone marrow mesenchymal stem cell-conditioned medium (BMSC-CM) on endometriosis (EMS) model in vivo and in vitro. METHODS: The animals were divided into two main groups, normal and EMS-induced mice. Normal and EMS-induced groups were injected with or without LA (250 mg/kg), LC (250 mg/kg), and BMSC-CM (a final volume of 100 µL of CM/mouse). At the end of the study, the level of total antioxidant capacity (TAC), nitric oxide (NO), and total oxidative status (TOS) were measured in plasma. Furthermore, immature oocytes were collected from two groups and cultured in a maturation medium. Subsequently, the rates of in vitro maturation, in vitro fertilization (IVF), and in vitro embryonic development were evaluated. RESULTS: The results revealed that administration of LA, LC, and BMSC-CM ameliorated the oxidative status through maintaining TAC and alleviating TOS and NO levels. More importantly, the maturation and fertilization rates, blastocyst development, and total blastocyst cell numbers significantly increased in LA, LC, and BMSC-CM-administrated groups compared to the control group. In both the normal and EMS groups, the highest IVF, cleavage, and blastocyst percentages were associated with BMSC-CM treatment (p < 0.05). CONCLUSION: Altogether, LA, LC, and BMSC-CM have therapeutic effects on impaired oocyte quality and promote subsequent development in vitro, probably through normalization of nitro-oxidative stress, thus offering potential alternatives to conventional therapies during assisted reproductive technologies for patients with EMS-associated sub/infertility.

Female Reproductive Aging and Oxidative Stress: Mesenchymal Stem Cell Conditioned Medium as a Promising Antioxidant.

The recent tendency to delay pregnancy has increased the incidence of age-related infertility, as female reproductive competence decreases with aging. Along with aging, a lowered capacity of antioxidant defense causes a loss of normal function in the ovaries and uterus due to oxidative damage. Therefore, advancements have been made in assisted reproduction to resolve infertility caused by reproductive aging and oxidative stress, following an emphasis on their use. The application of mesenchymal stem cells (MSCs) with intensive antioxidative properties has been extensively validated as a regenerative therapy, and proceeding from original cell therapy, the therapeutic effects of stem cell conditioned medium (CM) containing paracrine factors secreted during cell culture have been reported to be as effective as that of direct treatment of source cells. In this review, we summarized the current understanding of female reproductive aging and oxidative stress and present MSC-CM, which could be developed as a promising antioxidant intervention for assisted reproductive technology.

Effect of Passage Number of Conditioned Medium Collected from Equine Amniotic Fluid Mesenchymal Stem Cells: Porcine Oocyte Maturation and Embryo Development.

Oocyte in vitro maturation (IVM) is the most important first step in in vitro embryo production. One prerequisite for the success of IVM in oocytes is to provide a rich culture microenvironment that meets the nutritional needs of developing oocytes. We applied different equine amniotic fluid mesenchymal stem cell conditioned medium (eAFMSC-CM) from passages 7, 18, and 27 to porcine oocytes during IVM to determine its effects on oocyte development and subsequent embryo development, specifically. The eAFMSC-CM from passage 7 (eAFMSC-CMp7) has a considerable impact on 9 genes: BAX, BCL2, SOD2, NRF2, TNFAIP6, PTGS2, HAS2, Cx37, and Cx43, which are associated with cumulus cell mediated oocyte maturation. GSH levels and distribution of mitochondrial and cortical granules were significantly increased in oocytes incubated with eAFMSC-CMp7. In addition, catalase and superoxide dismutase activities were high after IVM 44 h with eAFMSC-CMp7. After in vitro fertilization, blastocyst quality was significantly increased in the eAFMSC-CMp7 group compared to control. Lastly, the antioxidant effect of eAFMSC-CMp7 substantially regulated the expression of apoptosis, pluripotency related genes and decreased autophagy activity in blastocysts. Taken together, this study demonstrated that the eAFMSC-CMp7 enhanced the cytoplasmic maturation of oocytes and subsequent embryonic development by generating high antioxidant activity.

Effects of the prevention of medication-related osteonecrosis of the jaw by local administration of a dental pulp stem cell-conditioned medium to the rat tooth extraction socket.

The onset of osteonecrosis of the jaw, which is a side effect of bisphosphonates, often develops after tooth extraction; measures for its prevention have not yet been established. While treatment with systemic administration of bone marrow stem cell-derived conditioned medium for medication-related osteonecrosis of the jaw (MRONJ) has been reported, its preventive effects have not been clarified yet, and the high degree of invasiveness of bone marrow fluid collection remains an issue. Therefore, we created a rat model of MRONJ using BP zoledronic acid, used a dental pulp stem cell-conditioned medium (DPSC-CM), which can be collected relatively easily, and locally applied it to the tooth extraction socket with atelocollagen and gelatin sponges. The preventive effect on the onset of MRONJ was subsequently examined. The results demonstrated that the bone exposure width of the extraction socket was reduced, and the mucosal covering was promoted in the atelocollagen + DPSC-CM group as compared with the other groups. Furthermore, histological results indicated a decrease in the number of empty bone lacunae, whereas immunohistochemical staining revealed the presence of many vascular endothelial growth factor (VEGF)-positive cells. Moreover, the results of the investigation of the sustained release of atelocollagen using VEGF indicated the release of VEGF over time. Our results suggest that local administration of DPSC-CM using atelocollagen may be a useful method for the prevention of MRONJ triggered by tooth extraction.

Adipose-derived stem cell-conditioned medium protects fibroblasts at different senescent degrees from UVB irradiation damages.

Adipose-derived stem cells (ADSCs) and their derivatives have aroused intense interest in fields of dermatological and aesthetic medicine. As a major component detected in ADSCs secretome, platelet-derived growth factor AA (PDGF-AA) has been reported mediating extracellular matrix deposition and remodeling, thus might contribute to its anti-aging effect. On the basis of establishing an experimental model that simulate actual skin aging by exposing HDFs to both intrinsic and extrinsic aging factors, we pretreated human dermal fibroblasts (HDFs) with ADSC-conditioned medium (ADSC-CM) before being irradiated, aiming at exploring preventive effects of ADSCs secretome against aging damages. 48 h after irradiation, we detected cellular proliferation; ß-galactosidase stain; mRNA expressions of MMP-1, MMP-9, and TIMP-1; and protein expressions of collagen I, collagen III, and elastin. Moreover, we detected related protein expression of PI3K/Akt signal pathway, which can be activated by PDGF-AA and was newly found to promote extracellular matrix protein synthesis. Concentration of PDGF-AA in the prepared ADSC-CM decreased over time and maintained excellent bioactivity at low temperature until the 11th week. ADSC-CM pretreatment can slightly or significantly improve cellular proliferative activity and reduce cellular senescence in irradiated HDFs. Besides, ADSC-CM pretreatment increased collagen I, collagen III, elastin, and TIMP-1 expressions but decreased MMP-1 and MMP-9 expressions both in irradiated and nonirradiated HDFs. ADSC-CM pretreatment significantly increased pAkt protein expression, and ECM protein expression greatly decreased in case of LY294002 application. The results were similar in three generations of HDFs, yet varied with different degrees. Generally, ADSC-CM we prepared demonstrates a certain degree of positive role in preventing HDFs from intrinsic and extrinsic aging damages and that PDGF-AA may contribute to making it become effective with some other components in ADSC-CM.

Improvement in infected wound healing in type 1 diabetic rat by the synergistic effect of photobiomodulation therapy and conditioned medium.

We investigated the effects of photobiomodulation therapy (PBMT) and conditioned medium (CM) of human bone marrow mesenchymal stem cells (hBM-MSC) individually and/or in combination on the stereological parameters and the expression of basic fibroblast growth factor (bFGF), hypoxia-inducible factor (HIF-1α), and stromal cell-derived factor-1α (SDF-1α) in a wound model infected with methicillin-resistant Staphylococcus aureus (MRSA) in diabetic rats. CM was provided by culturing hBM-MSCs. Type 1 diabetes mellitus (T1DM) was induced in 72 rats, divided into four groups, harboring 18 rats each: group 1 served as a control group, group 2 received PBMT, group 3 received CM, and group 4 received CM + PBMT. On days 4, 7, and 15, six animals from each group were euthanized and the skin samples were separated for stereology examination and gene expression analysis by real-time polymerase chain reaction. In the CM + PBMT, CM, and PBMT groups, significant decreases were induced in the number of neutrophils (1460 ± 93, 1854 ± 138, 1719 ± 248) and macrophages (539 ± 69, 804 ± 63, 912 ± 41), and significant increases in the number of fibroblasts (1073 ± 116, 836 ± 75, 912 ± 41) and angiogenesis (15 230 ± 516, 13 318 ± 1116, 14 041 ± 867), compared with those of the control group (2690 ± 371, 1139 ± 145, 566 ± 90, 12 585 ± 1219). Interestingly, the findings of the stereological examination in the CM + PBMT group were statistically more significant than those in the other groups. In the PBMT group, in most cases, the expression of bFGF, HIF-1α, and SDF-1α, on day 4 (27.7 ± 0.14, 28.8 ± 0.52, 27.5 ± 0.54) and day 7 (26.8 ± 1.4, 29.6 ± 1.4, 28.3 ± 1.2) were more significant than those in the control (day 4, 19.3 ± 0.42, 25.5 ± 0.08, 22.6 ± 0.04; day 7, 22.3 ± 0.22, 28.3 ± 0.59, 24.3 ± 0.19) and other treatment groups. The application of PBMT + CM induced anti-inflammatory and angiogenic activities, and hastened wound healing process in a T1 DM model of MRSA infected wound.

The effect of combined pulsed wave low-level laser therapy and mesenchymal stem cell-conditioned medium on the healing of an infected wound with methicillin-resistant Staphylococcal aureus in diabetic rats.

This study aims to investigate the combined effects of Pulsed wave low-level laser therapy (PW LLLT) and human bone marrow mesenchymal stem cell-conditioned medium (hBM-MSC-CM) on the microbial flora and tensiometrical properties of an infected wound model with methicillin-resistant staphylococcal aureus (MRSA) in an experimental model for Type 1 diabetes mellitus (TIDM). TIDM was induced in rats by streptozotocin (STZ). One full-thickness excision was made on the backs of the rats. Next, the rats were divided into the following groups: Group 1 was the control (placebo) group; Group 2 received hBM-MSCs-CM four times; Group 3 were laser PWLLLT (890 nm, 80 Hz, 0.2 J/cm2 ); and Group 4 received hBM-MSCs-CM +LASER. Wounds were infected with MRSA. Microbiological examinations were performed on days 4, 7, and 15. Tensiometerical examinations were carried out on the 15th day. One-way analysis of variance showed that laser and CM alone and/or in combination significantly increases the tensiomerical properties of the repaired wounds compared with control wounds. A combination of PW laser and CM was statistically more effective than other treated groups. Two-way analysis of variance showed that laser and CM alone and/or in combination significantly decreases the colony-forming units (CFUs) compared with the control group. The application of hBM-MSC-CM and PWlaser alone and/or together significantly accelerates the wound-healing process in MRSA-infected cutaneous wounds in TI DM in rats. Additionally, a combined application of hBM-MSC-CM and PWlaser demonstrates a synergistic effect on the wound-healing process in MRSA-infected cutaneous wounds in Type I DM rats.

Mesenchymal stem cell-conditioned medium accelerates wound healing with fewer scars.

Mesenchymal stem cells (MSCs) derived from umbilical cords (UC-MSCs) have been shown to enhance cutaneous wound healing by means of the paracrine activity. Fibroblasts are the primary cells involved in wound repair. The paracrine effects of UC-MSCs on dermal fibroblasts have not been fully explored in vitro or in vivo. Dermal fibroblasts were treated with conditioned media from UC-MSCs (UC-MSC-CM). In this model, UC-MSC-CM increased the proliferation and migration of dermal fibroblasts. Moreover, adult dermal fibroblasts transitioned into a phenotype with a low myofibroblast formation capacity, a decreased ratio of transforming growth factor-ß1,3 (TGF-ß1/3) and an increased ratio of matrix metalloproteinase/tissue inhibitor of metalloproteinases (MMP/TIMP). Additionally, UC-MSC-CM-treated wounds showed accelerated healing with fewer scars compared with control groups. These observations suggest that UC-MSC-CM may be a feasible strategy to promote cutaneous repair and a potential means to realise scarless healing.

Mesenchymal stem cell secreted platelet derived growth factor exerts a pro-migratory effect on resident Cardiac Atrial appendage Stem Cells.

Mesenchymal stem cells (MSCs) modulate cardiac healing after myocardial injury through the release of paracrine factors, but the exact mechanisms are still unknown. One possible mechanism is through mobilization of endogenous cardiac stem cells (CSCs). This study aimed to test the pro-migratory effect of MSC conditioned medium (MSC-CM) on endogenous CSCs from human cardiac tissue. By using a three-dimensional collagen assay, we found that MSC-CM improved migration of cells from human cardiac tissue. Cell counts, perimeter and area measurements were utilized to quantify migration effects. To examine whether resident stem cells were among the migrating cells, specific stem cell properties were investigated. The migrating cells displayed strong similarities with resident Cardiac Atrial appendage Stem Cells (CASCs), including a clonogenic potential of ~21.5% and expression of pluripotency associated genes like Oct-4, Nanog, c-Myc and Klf-4. Similar to CASCs, migrating cells demonstrated high aldehyde dehydrogenase activity and were able to differentiate towards cardiomyocytes. Receptor tyrosine kinase analysis and collagen assays performed with recombinant platelet derived growth factor (PDGF)-AA and Imatinib Mesylate, a PDGF receptor inhibitor, suggested a role for the PDGF-AA/PDGF receptor α axis in enhancing the migration process of CASCs. In conclusion, our findings demonstrate that factors present in MSC-CM improve migration of resident stem cells from human cardiac tissue. These data open doors towards future therapies in which MSC secreted factors, like PDGF-AA, can be utilized to enhance the recruitment of CASCs towards the site of myocardial injury.

Human umbilical cord blood mesenchymal stem cells reduce colitis in mice by activating NOD2 signaling to COX2.

BACKGROUND & AIMS: Decreased levels or function of nucleotide-binding oligomerization domain 2 (NOD2) are associated with Crohn's disease. NOD2 regulates intestinal inflammation, and also is expressed by human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs), to regulate their differentiation. We investigated whether NOD2 is required for the anti-inflammatory activities of MSCs in mice with colitis. METHODS: Colitis was induced in mice by administration of dextran sulfate sodium or trinitrobenzene sulfonic acid. Mice then were given intraperitoneal injections of NOD2-activated hUCB-MSCs; colon tissues and mesenteric lymph nodes were collected for histologic analyses. A bromodeoxyuridine assay was used to determine the ability of hUCB-MSCs to inhibit proliferation of human mononuclear cells in culture. RESULTS: Administration of hUCB-MSCs reduced the severity of colitis in mice. The anti-inflammatory effects of hUCB-MSCs were greatly increased by activation of NOD2 by its ligand, muramyl dipeptide (MDP). Administration of NOD2-activated hUCB-MSCs increased anti-inflammatory responses in colons of mice, such as production of interleukin (IL)-10 and infiltration by T regulatory cells, and reduced production of inflammatory cytokines. Proliferation of mononuclear cells was inhibited significantly by co-culture with hUCB-MSCs that had been stimulated with MDP. MDP induced prolonged production of prostaglandin (PG)E2 in hUCB-MSCs via the NOD2-RIP2 pathway, which suppressed proliferation of mononuclear cells derived from hUCB. PGE2 produced by hUCB-MSCs in response to MDP increased production of IL-10 and T regulatory cells. In mice, production of PGE2 by MSCs and subsequent production of IL-10 were required to reduce the severity of colitis. CONCLUSIONS: Activation of NOD2 is required for the ability of hUCB-MSCs to reduce the severity of colitis in mice. NOD2 signaling increases the ability of these cells to suppress mononuclear cell proliferation by inducing production of PGE2.

Therapeutic effect of human umbilical cord mesenchymal stem cells and their conditioned medium on LPS-induced endometritis in mice.

AIM: To explore the effect of human umbilical cord mesenchymal stem cells (hUC-MSCs) and their conditioned medium (MSC-CM) in repairing the endometritis mouse model in vivo. METHODS: Lipopolysaccharide (LPS) was used to induce acute inflammation in endometritis mouse model. Mice were treated in six groups: control group (PBS), model group (LPS), LPS+MSC-CM (6 h) group, LPS+MSC-CM (12 h) group, LPS+MSCs (6 h) group and LPS+MSCs (12 h) group. Morphological and histological changes of mouse uterus were observed, and mouse uterine inflammation index myeloperoxidase (MPO) and related immune index TNF-α, IL-6 and IL-1ß levels were detected by ELISA. RESULTS: There exist remarkable inflammatory response and an obvious increase in the value of MPO, TNF-α, IL-1ß and IL-6 in the endometritis mouse model compared with the control group. Morphological and histological appearances were relieved after treated with hUC-MSCs and MSC-CM. Besides, the value of MPO, TNF-α, IL-1ß and IL-6 showed different degrees of decline. In comparison with LPS+MSC-CM (12 h) and LPS+MSCs (12 h) group, there was significant decrease in inflammatory indicators in LPS+MSC-CM (6 h) and LPS+MSCs (6 h) group. CONCLUSIONS: Intrauterine infusion of hUC-MSCs and MSC-CM can alleviate LPS induced endometritis.

Stem cell-conditioned medium is a promising treatment for Alzheimer's disease.

BACKGROUND AND AIM: Alzheimer's disease (AD), a prevalent progressive neurodegenerative disease, is mainly characterized by dementia, memory loss, and cognitive disorder. Rising research was performed to develop pharmacological or non-pharmacological approaches to treat or improve AD complications. Mesenchymal stem cells (MSCs) are stromal cells that can self-renew and exhibit multilineage differentiation. Recent evidence suggested that some of the therapeutic effects of MSCs are mediated by the secreted paracrine factors. These paracrine factors, called MSC- conditioned medium (MSC-CM), may stimulate endogenous repair, promote angio- and artery genesis, and reduce apoptosis through paracrine mechanisms. The current study aims to systematically review the advantages of MSC-CM to the development of research and therapeutic concepts for AD management. MATERIAL AND METHODS: The present systematic review was performed using PubMed, Web of Science, and Scopus from April 2020 to May 2022 following the "Preferred Reporting Items for Systematic Reviews" (PRISMA) guidelines. The keywords, including "Conditioned medium OR Conditioned media OR Stem cell therapy" AND "Alzheimer's," was searched, and finally, 13 papers were extracted. RESULTS: The obtained data revealed that MSC-CMs might positively affect neurodegenerative diseases prognosis, especially AD, through various mechanisms, including a decrease in neuro-inflammation, reduction of oxidative stress and Aß formation, modulation of Microglia function and count, reduction of apoptosis, induction of synaptogenesis and neurogenesis. Also, the results showed that MSC-CM administration could significantly improve cognitive and memory function, increase the expression of neurotrophic factors, decrease the production of pro-inflammatory cytokines, improve mitochondrial function, reduce cytotoxicity, and increase neurotransmitter levels. CONCLUSION: While inhibiting the induction of neuroinflammation could be considered the first therapeutic effect of CMs, the prevention of apoptosis could be regarded as the most crucial effect of CMs on AD improvement.

The efficacy of topical adipose mesenchymal stem cell-conditioned medium versus framycetin gauze dressing in chronic plantar ulcer of leprosy: A randomized controlled trial.

Background Wound healing shows a unique interaction of several cells, growth factors and cytokines. The healing of chronic plantar ulcer of leprosy is influenced by various factors, one of which is the concentration of growth factors and cytokines related to the pathogenesis of impaired wound healing. Growth factors and cytokines can be found in the secretome of adipose mesenchymal stem cells. Aim To compare the effectiveness of topical adipose mesenchymal stem cell-conditioned medium and framycetin gauze dressing only on the healing of chronic plantar ulcer of leprosy. Methods In this randomised controlled trial, 32 patients with chronic plantar ulcer of leprosy were recruited. After detailed clinical and initial debridement, patients were randomised to two groups to receive either topical adipose mesenchymal stem cell-conditioned medium (n = 16) or framycetin gauze dressing only (n = 16) applied every three days for up to eight weeks, following which the ulcer size, adverse reactions and complications if any were monitored weekly. Results Healing percentage increased each week in all groups. Statistical differences between groups (P < 0.05) were observed from week 2 onwards for ulcer mean size reduction and from week 3 onwards for ulcer mean depth reduction. There were no adverse reactions or complications. Limitations Off-loading on subjects were not performed. Conclusion Adipose mesenchymal stem cell-conditioned medium is a potential therapeutic agent in the management of chronic plantar ulcer of leprosy.

Conditioned medium from bone marrow mesenchymal stem cells relieves spinal cord injury through suppression of Gal-3/NLRP3 and M1 microglia/macrophage polarization.

The therapeutic effects of bone marrow mesenchymal stem cells (BMSCs) and their conditioned media have been well-documented. This study focused on the effects of BMSC-conditioned medium (BMSCcm) on spinal cord injury (SCI). To study the effects of BMSCcm on rat motor function, inflammatory response, and M1/M2 macrophage/microglial polarization, SCI model rats were treated with BMSCcm and vectors for overexpression of galectin-3 (Gal-3) or NLR family pyrin domain containing 3 (NLRP3). Treatment with BMSCcm reduced the expression of Gal-3 and NLRP3, alleviated the inflammatory response, suppressed M1 microglia/macrophage polarization, and triggered M2 microglia/macrophage polarization in SCI model rats. Meanwhile, overexpression of Gal-3 or NLRP3 counteracted the suppressive effect of BMSCcm on SCI. Moreover, during BMSCcm treatment, overexpression of Gal-3 promoted the expression of NLRP3, whereas overexpression of NLRP3 had no significant effect on the expression of Gal-3. Additionally, the effects of BMSCcm on macrophage/microglial polarization and the underlying molecular mechanisms were observed in vitro. This study demonstrated that BMSCcm alleviates SCI by suppressing the expression of Gal-3 and NLRP3.

Conditioned medium from a human adipose-derived stem cell line ameliorates inflammation and fibrosis in a lung experimental model of idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis is a chronic, progressive parenchymal lung disease that results in fibrogenesis and the conditioned medium from adipose-derived mesenchymal stem cells (CM-ADSCs) has been shown to be efficacious in pulmonary fibrosis animal models. The aim of the present study is to evaluate the effect of CM-ADSCs on lung inflammation and fibrosis in a Bleomycin (BLM)-induced pulmonary fibrosis model. CM-ADSCs safety and toxicity were evaluated in Sprague Dawley rats and no adverse effects were observed. Six-week-old female C57BL/6J mice were employed in the BLM-induced pulmonary fibrosis model and were divided into four groups: Group 1 (Sham): animals were kept without BLM and treatment, Group 2 (Control): BLM with vehicle DMEM, Group 3: 10 µg/kg CM-ADSCs and Group 4: 100 µg/kg CM-ADSCs. Body weight, fibrosis and inflammation histological analyses, mRNA and protein pro-inflammatory cytokine, and total hydroxyproline content calculation were performed in all groups upon sacrifice. The 100 µg/kg CM-ADSCs showed a significant increase in mean body weight compared to Controls. CM-ADSCs doses resulted in the amelioration of fibrosis, as seen by Masson's Trichrome-staining, Ashcroft scoring, and Sirius red-staining. Compared to Controls, inflammation was also significantly reduced in CM-ADSCs-treated mice, with reduced F4/80 macrophage antigen staining, TNF-α mRNA and IL-6 and IL-10 protein levels. Total hydroxyproline content was found significantly reduced in both groups of CM-ADSCs-treated mice. Overall, our study shows that the CM-ADSCs is safe and efficient against pulmonary fibrosis, as it significantly reduced inflammation and fibrosis, with the larger dose of 100 µg/kg CM-ADSCs being the most efficient one.

In vitro effects of conditioned medium from bioreactor cultured human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) on skin-derived cell lines.

INTRODUCTION: When stem cells are grafted into tissues, they differentiate and form specialized cells. However, the proficiency of stem cells to endure and assimilate the host cell is dependent on various growth factors and cytokines. According to various studies, these factors are available in the spent media of harvested stem cells, which can be used for treatment in regenerative medicine and cosmetic products. There are differences in cytokine secretion depending on the culture environment, which are clarified in this paper. METHODS: Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) were cultured either in a bioreactor or in a flask. The conditioned medium from the hUC-MSC cultures in the flask and in the bioreactor was designated as "FM" and "BM", respectively. We assessed the effects of FM and BM on UVB-induced oxidative stress, anti-aging, and melanogenic properties. The amount of growth factors, cell viability, hyaluronic acid (HA), pro-collagen, and pro-melanin were quantitatively evaluated in the FM and BM treated groups. The induction of HA and collagen synthesis was measured in CCD-986SK cells. For melanogenesis, the effects of FM and BM on melanin content and tyrosinase activity were measured in SK-MEL-31 cells. RESULTS: In the present study, the secretion of growth factors, HA, and pro-collagen was significantly higher in the BM treatment, compared to that in the FM treatment. BM protected CCD-986SK cells against death from UVB induced oxidative stress. BM increased the promoter activity of the anti-oxidant genes SOD1, CAT, and GP; and downregulated the accelerating collagen decomposition gene, MMP-1, induced by UVB irradiation. In α-melanocyte-stimulating hormone (α-MSH) stimulated SK-MEL-31 cells, BM reduced melanin production and decreased the levels of MITF, tyrosinase, TRP-1, and TRP-2. These results suggest that BM could be used as a skin protection agent, because of its anti-apoptotic, anti-aging, and anti-melanogenic properties. This could be attributed to the differences in culturing methods; it is difficult to maintain the temperature and sterility in FM culture, when compared to that in the automated culturing conditions of the BM system. CONCLUSIONS: Collectively, our results indicate that using BM-conditioned hUC-MSC medium is very efficient process for producing raw materials for developing functional cosmetics.

Efficacy of stem cell-conditioned medium vs. platelet-rich plasma as an adjuvant to ablative fractional CO2 laser resurfacing for atrophic post-acne scars: a split-face clinical trial.

OBJECTIVES: To explore the impact of using topical stem cell-conditioned medium (SC-CM) after fractional carbon dioxide laser (FCL) vs. combined FCL and platelet-rich plasma (PRP) or FCL alone in treatment of atrophic acne scars. METHODS: Thirty-three patients were randomly divided into two split-face groups. Group I (n = 17) received FCL plus topical SC-CM on one side or FCL plus saline on the other. Group II (n = 16) received FCL plus topical PRP or SC-CM. All patients had three monthly sessions. Clinical assessment was done at each visit, with a final assessment after 3 months. Skin biopsies were obtained for histological and quantitative molecular analysis after treatment. RESULTS: No significant difference in clinical improvement of acne scars was observed between the FCL/SC-CM and FCL only sides (p = .63), while better and faster improvement was detected on FCL/PRP side compared to FCL/SC-CM side (p = .006). There was no significant difference in downtime or adverse effects between the treated sides in either group. Dermal collagen was increased and procollagen type I gene was upregulated in both FCL/PRP and FCL/SC-CM sides compared to FCL only sides (p = .001 and p = .041, respectively). CONCLUSIONS: Topical SC-CM could potentially enhance the efficacy of FCL. However, PRP seems to be a better alternative.

Hypoxic-conditioned medium from adipose tissue mesenchymal stem cells improved neuroinflammation through alternation of toll like receptor (TLR) 2 and TLR4 expression in model of Alzheimer's disease rats.

Microglia have a pivotal role to initiate immune responses in AD brains through toll-like receptors and induce neuroinflammation. Adipose tissue mesenchymal stem cells (ATSCs) secret many neurotrophic and anti-inflammatory factors called conditioned medium (CM). Many studies have demonstrated that CM of mesenchymal stem cells facilitate regeneration and attenuates inflammation in many disorders. To this purpose, the effect of ATSCs-conditioned medium (ATSC-CM) on brain inflammation and the role of toll-like receptors were investigated in this study. Seventy-two rats were randomly divided into 6 groups: control, sham, sham+ATSC-CM: 200µl ATSC-CM once a day intraperitoneally for 8 days, AD group injected the Aß1-40 intra-hippocampal, AD+ASC-CM, which was injected Aß1-40 intra-hippocampal and 200µl ATSC-CM once a day intraperitoneally for 8 days and AD+ rivastigmine: was injected Aß1-40 intra-hippocampal and received rivastigmine (0.6 mg/kg) orally once a day for 2 weeks. Memory and learning were measured by Morris water maze and novel object recognition tests. For detection of beta-amyloid plaque, Congo red staining was used, and neuronal survival was assessed by Nissl staining. Expression of TLR2 and TLR4 was measured by real-time PCR, and finally, to assess inflammation markers (IL-1ß and TNF-α) in the hippocampus, ELISA kits were used. In treatment group spatial and recognition memory significantly was improved. ATSC-CM administration decreased beta amyloid plaques and enhanced neuronal survival in AD brain rats. In addition, TLR2 and TLR4 expression decreased in treatment group. Results also showed that ATSC-CM reduced IL-1ß and TNF-α as inflammation markers. ATSC-CM improved memory deficit, decreased beta amyloids formation, increased neuron survival, and attenuated inflammation by reducing the expression of TLRs.

Mechanism of inhibiting proliferation of hepatocellular carcinoma Hepa1-6 cells by embryonic stem cell-conditioned medium.

The present study aimed to investigate the antiproliferative effect of embryonic stem cell-conditioned medium (ESC-CM) on the mouse liver cancer Hepa1-6 cells in vitro. Furthermore, in order to elucidate the underlying molecular mechanism, the microRNAs (miRNAs) in ESC-CM associated with the inhibition of Hepa1-6 proliferation were identified. Following the co-culture of ESC-CM and Hepa1-6 in Transwell chambers, the proliferation, cell cycle, apoptosis and associated protein expression were determined in Hepal-6 cells. Moreover, miRNA array analysis was employed to identify differentially expressed miRNAs. Based on the differentially expressed miRNAs, the target genes and potential associated signaling pathways were determined. Finally, RT-qPCR was conducted to confirm the above results. The ESC-CM inhibited Hepal-6 cell proliferation and increased the percentage of cells at G1 phase and decreased the percentage of cells at the G2/M phase of the cell cycle. The expression of cyclin D1/cyclin-dependent kinase (CDK)4/CDK6 was decreased following co-culture, with no effect on cell apoptosis. Six significantly regulated miRNAs were identified and 423 putative target genes of these regulated miRNAs were predicted. Gene ontology analysis revealed the putative target genes to be associated with the 'DNA replication (GO: 0006260)' GO term, 'apoptosis' and 'signal transduction'. The Kyoto Encyclopedia of Genes and Genomes analysis indicated that deregulated miRNAs were enriched in the Wnt signaling (KEGG entry: Map 04310) and Hippo signaling pathways (KEGG entry: Map 04390), pathways associated with cancer. Overall, the present study demonstrated the inhibition of Hepa1-6 cell line proliferation upon treatment with ESC-CM, by decreasing cell cycle-associated protein cyclin D1/CDK4/CDK6 expression and arresting cells in G1 phase of the cell cycle, with no effect on cell apoptosis. Furthermore, the inhibition of proliferation by ESC-CM may be mediated by miRNAs that affect cell cycle-associated mRNAs and the Wnt signaling pathway.