Analysis of the correlation between serum Klotho and FeNO: a cross-sectional study from NHANES (2007-2012).

BACKGROUND: Klotho is an anti-aging protein that has multiple functions and may play a key role in the pathogenesis and progression of chronic respiratory diseases such as chronic obstructive pulmonary disease (COPD). Fractional Exhaled Nitric Oxide (FeNO) is a non-invasive and novel biomarker that has the advantages of being simple, fast and reproducible. It can effectively assess the degree of airway inflammation in diseases such as asthma and COPD. Despite these insights, the relationship between serum Klotho levels and FeNO has not been explored yet. METHODS: Leveraging data from the National Health and Nutrition Examination Survey (NHANES) spanning 2007 to 2012, we investigated the correlation between FeNO and serum Klotho levels. This association was scrutinized both as continuous variables and within quartile distributions, utilizing the Kruskal-Wallis H test. The correlation between the two variables was assessed through Spearman rank analysis. Employing survey weight-adjusted linear regression models, we gauged the strength of these associations. RESULTS: This study included 6,527 participants with a median FeNO level of 14.5 parts per billion (ppb). We found that FeNO levels varied significantly across different quartiles of Klotho protein (H = 7.985, P = 0.046). We also found a significant positive correlation between serum Klotho levels and FeNO levels in the whole population (Spearman's rho = 0.029, P = 0.019). This correlation remained significant after adjusting for covariates such as age, gender, lung function, smoking status, alcohol use, BMI, cardiovascular disease (including hypertension, heart failure, coronary heart disease, and myocardial infarction), diabetes, inflammatory markers, serum vitamin D level and BUN (P < 0.05 for all). Furthermore, this correlation was stronger at the high (K3) and super high (K4) levels of Klotho than at the low (K1) and medium (K2) levels (ß = 1.979 ppb and ß = 1.993 ppb for K3 and K4 vs. K1, respectively; 95% CI: 0.497 ~ 2.953 and 95% CI: 0.129 ~ 2.827, respectively; P = 0.007 and P = 0.032, respectively). The ß coefficient for serum Klotho was 0.002 ppb/pg/ml. CONCLUSIONS: Our study illuminates a positive correlation between serum Klotho levels and FeNO. Further study is needed to verify the causality of this association and elucidate the underlying mechanisms.

Natural compounds protect against the pathogenesis of osteoarthritis by mediating the NRF2/ARE signaling.

Osteoarthritis (OA), a chronic joint cartilage disease, is characterized by the imbalanced homeostasis between anabolism and catabolism. Oxidative stress contributes to inflammatory responses, extracellular matrix (ECM) degradation, and chondrocyte apoptosis and promotes the pathogenesis of OA. Nuclear factor erythroid 2-related factor 2 (NRF2) is a central regulator of intracellular redox homeostasis. Activation of the NRF2/ARE signaling may effectively suppress oxidative stress, attenuate ECM degradation, and inhibit chondrocyte apoptosis. Increasing evidence suggests that the NRF2/ARE signaling has become a potential target for the therapeutic management of OA. Natural compounds, such as polyphenols and terpenoids, have been explored to protect against OA cartilage degeneration by activating the NRF2/ARE pathway. Specifically, flavonoids may function as NRF2 activators and exhibit chondroprotective activity. In conclusion, natural compounds provide rich resources to explore the therapeutic management of OA by activating NRF2/ARE signaling.

Role of IL-22 in intestinal microenvironment and potential targeted therapy through diet.

IL-22 is a type 2 receptor cytokine in IL-10 family. IL-22 is usually secreted by innate and adaptive immune cells and takes its effects on non-hematopoietic cells. Through activate STAT3 pathway, IL-22 plays an important role in infection clearance and tissue regeneration, which is critical for barrier integrate and homeostasis. Abnormal activation of IL-22 signal was observed in inflammation diseases, autoimmune diseases, and cancers. We review the recent discoveries about the mechanism and regulation of IL-22 signal pathway from the perspective of intestinal micro-environment. Diet-based IL-22 target therapeutic strategies and their potential clinical significance will also be discussed.

mTORC2 Facilitates Liver Regeneration Through Sphingolipid-Induced PPAR-α-Fatty Acid Oxidation.

BACKGROUND & AIMS: During liver regeneration after partial hepatectomy, the function and metabolic pathways governing transient lipid droplet accumulation in hepatocytes remain obscure. Mammalian target of rapamycin 2 (mTORC2) facilitates de novo synthesis of hepatic lipids. Under normal conditions and in tumorigenesis, decreased levels of triglyceride (TG) and fatty acids (FAs) are observed in the mTORC2-deficient liver. However, during liver regeneration, their levels increase in the absence of mTORC2. METHODS: Rictor liver-specific knockout and control mice underwent partial hepatectomy, followed by measurement of TG and FA contents during liver regeneration. FA metabolism was evaluated by analyzing the expression of FA metabolism-related genes and proteins. Intraperitoneal injection of the peroxisome proliferator-activated receptor α (PPAR-α) agonist, p53 inhibitor, and protein kinase B (AKT) activator was performed to verify the regulatory pathways involved. Lipid mass spectrometry was performed to identify the potential PPAR-α activators. RESULTS: The expression of FA metabolism-related genes and proteins suggested that FAs are mainly transported into hepatocytes during liver regeneration. The PPAR-α pathway is down-regulated significantly in the mTORC2-deficient liver, resulting in the accumulation of TGs. The PPAR-α agonist WY-14643 rescued deficient liver regeneration and survival in mTORC2-deficient mice. Furthermore, lipidomic analysis suggested that mTORC2 deficiency substantially reduced glucosylceramide (GluCer) content. GluCer activated PPAR-α. GluCer treatment in vivo restored the regenerative ability and survival rates in the mTORC2-deficient group. CONCLUSIONS: Our data suggest that FAs are mainly transported into hepatocytes during liver regeneration, and their metabolism is facilitated by mTORC2 through the GluCer-PPAR-α pathway, thereby establishing a novel role for mTORC2 in lipid metabolism.

Dimethyl itaconate alleviates the pyroptosis of macrophages through oxidative stress.

Macrophages are involved in the pathophysiology of many diseases as critical cells of the innate immune system. Pyroptosis is a form of macrophage death that induces cytokinesis of phagocytic substances in the macrophages, thereby defending against infection. Dimethyl itaconate (DI) is an analog of itaconic acid with anti-inflammatory effects. However, the effect of dimethyl itaconate on macrophage pyroptosis has not been elucidated clearly. Thus, the present study aimed to analyze the effect of DI treatment on a macrophage pyroptosis model (Lipopolysaccharide, LPS + Adenosine Triphosphate, ATP). The results showed that 0.25 mM DI ameliorated macrophage pyroptosis and downregulated interleukin (IL)-1ß expression. Then, real-time quantitative polymerase chain reaction (RT-qPCR) was used to confirm the result of RNA-sequencing of the upregulated oxidative stress-related genes (Gclc and Gss) and downregulated inflammation-related genes (IL-12ß and IL-1ß). In addition, Gene Ontology (GO) enrichment analysis showed that differential genes were associated with transcript levels and DNA replication. Kyoto encyclopedia of genes and genomes (KEGG) enrichment showed that signaling pathways, such as tumor necrosis factor (TNF), Jak, Toll-like receptor and IL-17, were altered after DI treatment. N-acetyl-L-cysteine (NAC) reversed the DI effect on the LPS + ATP-induced macrophage pyroptosis and upregulated the IL-1ß expression. Oxidative stress-related protein Nrf2 is involved in the DI regulation of macrophage pyroptosis. Taken together, these findings suggested that DI alleviates the pyroptosis of macrophages through oxidative stress.

Macrophages enhance lipopolysaccharide induced apoptosis via Ang1 and NF-κB pathways in human umbilical vein endothelial cells.

Lipopolysaccharide (LPS) could induce apoptosis and dysfunction of endothelial cells. We aimed to reveal the effects of macrophages on cell proliferation and apoptosis in LPS induced human umbilical vein endothelial cells (HUVECs). THP-1 derived macrophages and HUVECs were co-cultured in the presence of LPS. Cell viability was measured by Cell Counting Kit-8 and apoptosis was analyzed by flow cytometry. Expression of Ang1, the NF-κB component p65 was evaluated by western blot and quantitative PCR. Small interfering RNAs (siRNAs) were used to knockdown the expression of proinflammatory cytokines and p65 in HUVECs. Plasmid transfection-mediated overexpression of Ang1 was employed to see its effects on cell proliferation and apoptosis in HUVECs. Macrophages enhanced LPS-induced proliferation impairments and apoptosis in HUVECs, which could be attenuated by siRNA-mediated knockdown of cytokines TNF-α, IL-1ß, IL-6 and IL-12p70 in macrophages. The dysfunction of HUVECs was tightly associated with reduced Ang1 expression and increased phosphorylated p65 (p-65). Overexpression of Ang1 in HUVECs significantly decreased p-p65, suggesting negatively regulation of p-p65 by Ang1. Overexpression of Ang1, adding recombinant Ang1 or silencing of p65 substantially attenuated the dysfunction of HUVECs in terms of cell proliferation and apoptosis. In conclusions, THP-1-derived macrophages enhance LPS induced dysfunction of HUVECs via Ang1 and NF-κB pathways, suggesting new therapeutic targets for sepsis.

Upregulation of miR-92a contributes to blocking goblet cell metaplasia by targeting MUC5AC in asthma.

As a chronic airway disease, asthma has two characteristics, tissue remodeling and airway inflammation. This research focused on miR-92a to explore how it works in asthma. We revealed that the expressions of miR-92a were decreased in both serum and lung tissues from ovalbumin-induced asthma mouse. Bioinformatics analysis, quantitative polymerase chain reaction (qPCR) and dual luciferase assay revealed that miR-92a targets MUC5AC, which was linked to mucus hypersecretion in the pulmonary tracts. By injecting miR-92a-mimics into the trachea, both the airway hyper-reactivity and airway inflammation can be alleviated in an asthma mouse model which is induced by ovalbumin. Moreover, the goblet cell phenotype of asthmatic mice is significantly reduced by the action of miR-92a. Furthermore, miR-92a blocked interleukin (IL)-13-induced MUC5AC luciferase activity in 16HBE. Together, upregulation of miR-92a expression in asthmatic mice plays a role in blocking goblet cell metaplasia by targeting MUC5AC, and thus in the treatment of chronic airway diseases, miR-92a can prevent epithelial remodeling, which is a reasonable method.

Association between lncRNA GAS5, MEG3, and PCAT-1 Polymorphisms and Cancer Risk: A Meta-Analysis.

PURPOSE: Long noncoding RNAs (lncRNAs) have been widely studied, and single nucleotide polymorphisms (SNPs) in lncRNAs are considered to be genetic factors that influence cancer susceptibility. The lncRNA GAS5, MEG3, and PCAT-1 polymorphisms are shown to be possibly associated with cancer risk. The aim of this meta-analysis was to systematically evaluate this association. METHODS: Studies were selected from PubMed, Web of Science, Embase, Google Scholar, Cochrane Library, the Chinese National Knowledge Infrastructure (CNKI), and the Chinese Biomedical Literature Database (CBM) through inclusion and exclusion criteria. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using the random-effects model or fixed-effects model to assess the association between lncRNA polymorphisms and cancer susceptibility. Metaregression and publication bias analyses were also conducted. All analyses were performed using the Stata 12.0 software. RESULTS: Sixteen articles (covering 13750 cases and 17194 controls) were included in this meta-analysis. A significant association between SNP rs145204276 and gastric cancer risk was observed (del vs. ins: OR = 0.79, 95%CI = 0.72-0.86; del/del vs. ins/ins+del/ins: OR = 0.74, 95%CI = 0.59-0.91; del/ins vs. ins/ins: OR = 0.84, 95%CI = 0.67-1.05). For rs16901904, a decreased cancer risk was observed in three genetic models (C vs. T: OR = 0.79, 95%CI = 0.70-0.90; CC vs. CT+TT: OR = 0.49, 95%CI = 0.37-0.65; CC vs. TT: OR = 0.49, 95%CI = 0.37-0.66). No statistical significance was found in the metaregression analysis. For all of the included SNPs, no publication bias was found in all genotype models. CONCLUSIONS: The rs145204276 SNP in lncRNA GAS5 is likely to be associated with gastric cancer risk, whereas the rs16901904 SNP in lncRNA PCAT-1 bears association with a decreased cancer risk.

[Paeoniflorin improves the permeability of cardiac microvascular endothelial cells by regulating Src/vascular endothelial-cadherin pathway].

OBJECTIVE: To investigate the effect and mechanism of paeoniflorin on the permeability of cardiac microvascular endothelial cells (CMECs) in sepsis. METHODS: Primary rat CMECs were isolated and cultured in vitro, and the cells in the logarithmic growth phase were used for experiments. Tetramethylazozolium colorimetry (MTT) was used to screen the safe and effective concentrations of paeoniflorin at 10, 20, and 40 µmol/L. The cells were divided into blank control group, lipopolysaccharide (LPS) group and low, medium and high concentration paeoniflorin pretreatment group. The cells in the blank control group were cultured in complete medium; the cells in the LPS group were challenged with LPS (1 mg/L) in complete medium; and the cells in the paeoniflorin pretreatment groups were pretreated with 10, 20, and 40 µmol/L paeoniflorin at 4 hours before LPS stimulation. The cells in each group were further cultured for 24 hours after LPS stimulation. The horseradish peroxidase (HRP) method was used to detect the permeability of rat CMECs. The enzyme-linked immunosorbent assay (ELISA) was used to detect the CXC chemokine ligand (CXCL1, CXCL2) levels in the cell supernatant. The real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of CXCL1 and CXCL2 in the cells. Western Blot was used to detect phosphorylated Src (p-Src), vascular endothelial-cadherin (VE-cadherin) and phosphorylated mitogen activated protein kinase (p-MAPK). RESULTS: Compared with the blank control group, the permeability of rat CMECs in the LPS group was significantly increased. The cell permeability was improved to some extent after paeoniflorin pretreatment at different concentrations, and the improvement was most obvious in the 40 µmol/L paeoniflorin group, with statistically significant difference as compared with the LPS group (A value: 1.61±0.07 vs. 2.13±0.06, P < 0.01). ELISA results showed that there were moderate amounts of CXCL1 and CXCL2 in the cell supernatant of rat CMECs in the blank control group. However, the secretion of CXCL1 and CXCL2 in the cell supernatant was increased significantly under the induction of LPS. After pretreatment with paeoniflorin at different concentrations, the secretion of CXCL1 and CXCL2 in the cell supernatant was significantly reduced. The most obvious inhibitory effect on CXCL1 was 40 µmol/L paeoniflorin, and the most obvious inhibition on CXCL2 was 20 µmol/L paeoniflorin, the differences were statistically significant as compared with the LPS group [CXCL1 (ng/L): 337.51±68.04 vs. 829.86±65.06, CXCL2 (ng/L): 4.48±0.11 vs. 9.41±0.70, both P < 0.01]. RT-qPCR results showed that the mRNA expressions of CXCL1 and CXCL2 in the rat CMECs were consistent with the ELISA results. LPS could increase mRNA expressions of CXCL1 and CXCL2 in the rat CMECs, and pretreatment with different concentrations of paeoniflorin could significantly reduce the mRNA expressions of CXCL1 and CXCL2. The 40 µmol/L paeoniflorin had the best inhibitory effect on CXCL1 mRNA expression, and the 20 µmol/L paeoniflorin had the best inhibitory effect on CXCL2 mRNA expression, the differences were statistically significant as compared with the LPS group [CXCL1 mRNA (2-ΔΔCt): 0.543±0.004 vs. 0.812±0.089, CXCL2 mRNA (2-ΔΔCt): 10.52±0.71 vs. 17.68±1.09, both P < 0.01]. Western Blot results showed that moderate amounts of p-Src, VE-cadherin and p-MAPK proteins were expressed in the rat CMECs in the blank control group. After LPS stimulation, the expressions of p-Src and p-MAPK proteins were increased significantly, while the expression of VE-cadherin protein was decreased significantly. After pretreatment with different concentrations of paeoniflorin, the expressions of p-Src and p-MAPK proteins in the cells were decreased to varying degrees, while the expression of VE-cadherin protein was increased, and 40 µmol/L paeoniflorin had the most obvious effect, the differences were statistically significant as compared with the LPS group [p-Src protein (p-Src/GAPDH): 1.02±0.09 vs. 1.29±0.05, p-MAPK proteins (p-MAPK/GAPDH): 0.24±0.02 vs. 0.62±0.02, VE-cadherin protein (VE-cadherin/GAPDH): 0.64±0.03 vs. 0.31±0.02, all P < 0.01]. CONCLUSIONS: Paeoniflorin can regulate the Src/VE-cadherin pathway in CMECs, inhibit the expression and secretion of inflammation-related proteins and chemokines, and improve the cell permeability of CMECs induced by LPS.

VCAM-1+ placenta chorionic villi-derived mesenchymal stem cells display potent pro-angiogenic activity.

INTRODUCTION: Mesenchymal stem cells (MSCs) represent a heterogeneous cell population that is promising for regenerative medicine. The present study was designed to assess whether VCAM-1 can be used as a marker of MSC subpopulation with superior angiogenic potential. METHODS: MSCs were isolated from placenta chorionic villi (CV). The VCAM-1(+/-) CV-MSCs population were separated by Flow Cytometry and subjected to a comparative analysis for their angiogenic properties including angiogenic genes expression, vasculo-angiogenic abilities on Matrigel in vitro and in vivo, angiogenic paracrine activities, cytokine array, and therapeutic angiogenesis in vascular ischemic diseases. RESULTS: Angiogenic genes, including HGF, ANG, IL8, IL6, VEGF-A, TGFß, MMP2 and bFGF, were up-regulated in VCAM-1(+)CV-MSCs. Consistently, angiogenic cytokines especially HGF, IL8, angiogenin, angiopoitin-2, µPAR, CXCL1, IL-1ß, IL-1α, CSF2, CSF3, MCP-3, CTACK, and OPG were found to be significantly increased in VCAM-1(+) CV-MSCs. Moreover, VCAM-1(+)CV-MSCs showed remarkable vasculo-angiogenic abilities by angiogenesis analysis with Matrigel in vitro and in vivo and the conditioned medium of VCAM-1(+) CV-MSCs exerted markedly pro-proliferative and pro-migratory effects on endothelial cells compared to VCAM-1(-)CV-MSCs. Finally, transplantation of VCAM-1(+)CV-MSCs into the ischemic hind limb of BALB/c nude mice resulted in a significantly functional improvement in comparison with VCAM-1(-)CV-MSCs transplantation. CONCLUSIONS: VCAM-1(+)CV-MSCs possessed a favorable angiogenic paracrine activity and displayed therapeutic efficacy on hindlimb ischemia. Our results suggested that VCAM-1(+)CV-MSCs may represent an important subpopulation of MSC for efficient therapeutic angiogenesis.

Human mesenchymal stem cells possess different biological characteristics but do not change their therapeutic potential when cultured in serum free medium.

INTRODUCTION: Mesenchymal stem cells (MSCs) are widely investigated in clinical researches to treat various diseases. Classic culture medium for MSCs, even for clinical use, contains fetal bovine serum. The serum-containing medium (SCM) seems a major obstacle for MSCs-related therapies due to the risk of contamination of infectious pathogens. Some studies showed that MSCs could be expanded in serum free medium (SFM); however, whether SFM would change the biological characteristics and safety issues of MSCs has not been well answered. METHODS: Human umbilical cord mesenchymal stem cells (hUC-MSCs) were cultured in a chemical defined serum free medium. Growth, multipotency, surface antigen expression, telomerase, immunosuppressive ability, gene expression profile and genomic stability of hUC-MSCs cultured in SFM and SCM were analyzed and compared side by side. RESULTS: hUC-MSCs propagated more slowly and senesce ultimately in SFM. SFM-expanded hUC-MSCs were different from SCM-expanded hUC-MSCs in growth rate, telomerase, gene expression profile. However, SFM-expanded hUC-MSCs maintained multipotency and the profile of surface antigen which were used to define human MSCs. Both SFM- and SCM-expanded hUC-MSCs gained copy number variation (CNV) in long-term in vitro culture. CONCLUSION: hUC-MCSs could be expanded in SFM safely to obtain enough cells for clinical application, meeting the basic criteria for human mesenchymal stem cells. hUC-MSCs cultured in SFM were distinct from hUC-MSCs cultured in SCM, yet they remained therapeutic potentials for future regenerative medicine.

[Long term in-vitro expansion reduces immune modulation function of placental chorionic villi mesenchymal stem cells].

The main aim of this study was to investigate the biological activities and immune modulation changes of chorionic villi mesenchymal stem cells (CV-MSC) after long term culture. The morphology of the CV-MSC of passage 3 and passage 9 were observed by microscopy, and their phenotypes were detected by flow cytometry. CV-MSC of passage 3 and 9 were co-cultured with PHA-stimulated PBMNC, and IFN-γ concentration in culture medium was detected by ELISA. The mRNA expression of COX-2, HGF and HLA-G in CV-MSC were detected by real-time PCR. The results showed that after long term culture, the CV-MSC kept the MSC morphology and most of the phenotypes including CD31, CD34, CD44, CD45, CD62L, CD73, CD90, CD105, CD117, CD151, CD235a, CD271 and HLA-DR, while the CD49d was significantly up-regulated. Immune modulation ability of CV-MSC was reduced and the mRNA expression of COX-2 and HGF was down regulated after long term culture, but the expression of HLA-G did not found to be obvious change. It is concluded that the long term in vitro expansion changes the expression of CD49d and reduces immune modulation of CV-MSC.

[IL-1ß promotes the hematopoietic support of human umbilical cord mesenchymal stem cells].

This study was aimed to investigate the effect of IL-1ß on hematopoietic support of human umbilical cord mesenchymal stem cells (hUC-MSC). 2×10(6) hUC-MSC were seeded in 75 cm(2) flasks, after adherence to wall for 2 h, 10 ng/ml IL-1ß was added in hUC-MSC supernatant and cultured for 36 h, then the culture supernatants and cells were harvested. The effect of conditioned medium with/without IL-1ß on CD34(+) cell hematopoietic support was observed, mRNA expression changes of hUC-MSC cultured in medium with/without IL-1ß were monitored by real time PCR, the differences in hematopoiesis-related factors were detected by ELISA. The results showed that the conditioned culture medium of hUC-MSC with IL-1ß enhanced the ability to form colony of CD34(+) cells, especially CFU-G and CFU-GM in vitro; IL-1ß promoted the mRNA expression of GM-CSF, G-CSF, IL-6 on MSC; IL-1ß also promoted the secretion of GM-CSF, G-CSF, and IL-6 protein from hUC-MSC. It is concluded that IL-1ß enhances hematopoietic support capacity especially, capability of MSC to myeloid differentiation.

MicroRNA-124 suppresses breast cancer cell growth and motility by targeting CD151.

BACKGROUND: CD151 is highly expressed in breast cancer cells and has been shown to accelerate breast cancer by enhancing cell growth and motility, but its regulation is poorly understood. To explore post-translation regulation of CD151, for example microRNAs, will be of great importance to claim the mechanism. METHODS: A luciferase reporter assay was used to determine whether CD151 was a target of miR-124. The levels of CD151 mRNA were detected by real-time PCR and CD151 protein expression was measured by western blot and flow cytometry. The effects of miR-124 expression on growth, apoptosis, cell cycle and motility of breast cancer cells were determined. RESULTS: We discovered that miR-124 directly targets the 3' untranslated region (3'-UTR) of CD151 mRNAs and suppresses its mRNA expression and protein translation. Both siRNA of CD151 and miR-124 mimics could significantly inhibit proliferation of breast cancer cell lines via cell cycle arrest but does not induce apoptosis. Meanwhile, miR-124 mimics significantly inhibited the motility of breast cancer cells. CONCLUSION: miR-124 plays a critical role in inhibiting the invasive and metastatic potential of breast cancer cells, probably by directly targeting the CD151 genes. Our findings highlight an important role of miR-124 in the regulation of invasion and metastasis by breast cancer cells and suggest a potential application for miR-124 in breast cancer treatment.

[Isolation and biological characteristics of mesenchymal stem cells derived from human placenta decidua basalis].

Comparing to bone marrow mesenchymal stem cells (MSCs), placenta-derived MSCs have the advantages of adequate sources, low immunogenicity, little risk of viral contamination, and no ethical controversy, and thus possess a better prospect for clinical application. Placental tissue not only includes chorionic and amniotic, but also contains decidua basalis which locate in the maternal placenta surface. The biological characteristics of MSCs isolated from decidua basalis have not been well studied. This study was aimed to investigate the biologic characteristics of placenta decidua basalis-derived MSC from placenta decidua basalis (DB) by enzymatic digestion. Short tandem repeats (STR) test was used to identify the cells derived from the maternal placenta surface. Growth rate of decidua basalis mesenchymal stem cells (DB-MSC) was measured by MTT. Cell cycle and cell phenotype were detected by flow cytometry. Inducing differentiation was used to evaluate multipotency of DB-MSC. For testing the immunosuppression of DB-MSC, they were co-cultured with peripheral blood mononuclear cells (PBMNC) stimulated by phytohemagglutinin (PHA) and then IFN-γ in the co-cultured media was quantified by ELISA. The results showed that the cells were derived from the maternal placenta by STR analysis. DB-MSC showed typical fibroblast morphology in the culture and were positive for the MSC surface markers: CD90, CD73, CD105, CD44 and negative for CD45, CD11b, and CD34. DB-MSC underwent osteogenic, adipogenic and chondrogenic differentiation in inducing medium. DB-MSC could inhibit the secretion of IFN-γ by PBMNC. It is concluded that the cells are isolated from placenta decidua basalis and possess the basic characteristics of MSC. DB-MSC can be an important maternal autologous MSC and may be a safe and effective treatment for immune system diseases, which makes the DB-MSC as an important source of autologous MSC from mother. DB-MSC can be safely for the treatment of the mother's immune system diseases.

[Supportive effects of conditioned culture media of human umbilical cord mesenchymal stem cells on hematopoiesis in vitro].

This study was aimed to explore whether the conditioned culture medium of human umbilical cord-derived mesenchymal stem cells (hUC-MSC) has supportive effects on hematopoiesis in vitro. hUC-MSC were cultured in 75 cm(2) culture flasks at a concentration of 2×10(6) cells per flask. After 48 h, the conditioned culture medium was harvested. CD34(+) cells were isolated with the human cord blood CD34 positive selection kit. The CD34(+) cells were plated in three different culture systems: the culture supernatant from hUC-MSC added into incomplete methylcellulose without recombinant human cytokines as conditioned culture medium; the complete methylcellulose medium with recombinant human cytokines as positive control medium; incomplete methylcellulose adding DMEM/F12 with 10% FBS instead of conditioned culture medium as the negative control medium. After 14 days of culture, colonies containing ≥ 50 cells were scored and types of colonies were classified under inverted microscope. The immunophenotypes of cells which were collected from the colonies were detected by flow cytometry. The results showed that conditioned culture medium of hUC-MSC supported the differentiation of CD34(+) cells into CFU-G (47.67 ± 0.58), CFU-GM (48.67 ± 4.73) and CFU-M (3.00 ± 2.00) in vitro, while the CFU-E, BFU-E or CFU-GEMM were absent. Comparatively, in the positive control medium all kinds of CFU were observed. Interestingly, the percentage of CD45(+)cells of CFU in conditioned culture medium (97.43 ± 2.15)% was more than CD45(+)cells in positive control medium (39.69 ± 0.96)% (P < 0.05). It is concluded that the conditioned culture medium of hUC-MSC has been confirmed to have ability to support hematopoiesis separately in vitro. Besides, it enhances the differentiation of CD34(+) cells into myeloid cells except cells of erythroid lineage.

[Comparative study of biological characteristics of human umbilical cord and placental chorionic villous mesenchymal stem cells].

Because advantage of tissue origin and proliferation potential, the umbilical cord-derived mesenchymal stem cells (UC-MSC) and placental chorionic villous-derived mesenchymal stem cells (CV-MSC) have clinical application potential, as compared with bone marrow MSC. But whether the differences of biological characteristics exist between UC-MSC and CV-MSC, which deserve to be further explored. This study was purposed to compare the biological characteristics of UC-MSC and CV-MSC. The placental and umbilical cord were cleaned by using the sterile physiological salt, the UC-MSC and CV-MSC were separated by enzyme digestion. Short tandem repeat (STR) analysis was used to detect whether the MSC obtained from fetal tissue. MTT method was used to detect proliferation of MSC. Flow cytometry was applied to analyze cell phenotype. The different differential medium was used to detect their multi-directional differentiation capacity. After the MSC and PHA-stimulated peripheral blood mononuclear cells were co-cultured, the γ-interferon (IFN-γ) levels of the co-culture supernatant were detected using the ELISA. The results showed that these MSC were derived from fetal tissue by STR analysis. They were adherent cells with typical fibroblast morphology. Cells expressed the MSC surface markers CD90, CD73 and CD105 and CD44, not expressed CD45 and of CD11b and CD34.These cells could differentiate into osteoblasts and adipoblasts under culture with different conditioned medium, but in the adipogenic differentiation of CV-MSC, the larger lipid droplets appeared. It is concluded that these cells are obtained MSC. These MSC can inhibit peripheral blood mononuclear cells stimulated by PHA to secrete IFN-γ, and the the CV-MSC have a stronger suppression capacity, which makes the CV-MSC to have a greater advantage in the treatment of autoimmune diseases.

Intrapulmonary delivery of human umbilical cord mesenchymal stem cells attenuates acute lung injury by expanding CD4+CD25+ Forkhead Boxp3 (FOXP3)+ regulatory T cells and balancing anti- and pro-inflammatory factors.

BACKGROUND: Systemic and local inflammatory processes play key, mainly detrimental roles in the pathophysiology of acute lung injury (ALI). The present study was designed to determine whether human umbilical cord mesenchymal stem cells (UCMSC) are able to act on CD4(+)CD25(+) Foxp3(+)Treg cells and lead to an improvement in ALI. METHODS: Mice were administered intratracheally endotoxin (lipopolysaccharide [LPS]) and received intrapulmonary 1×10(6) UCMSC 4 hours after challenge. The CD4(+)CD25(+) Foxp3(+)Treg, survival time, body weight, histology and lung injury scores were assessed after transplantation of UCMSC. In addition, anti-inflammatory factor IL10 and pro-inflammatory mediators production including tumor necrosis factor-a (TNF-α), macrophage inflammatory protein-2(MIP-2) and interferon-γ (IFN-γ) were detected. RESULTS: Transplantation of UCMSC resulted in significant increase in the level of CD4(+)CD25(+) Foxp3(+)Treg in ALI. Increased level of anti-inflammatory factor IL-10 and reduced levels of TNF-α, MIP-2 and IFN-γ were simultaneously observed in ALI in comparison with control mice. CONCLUSION: Our data demonstrate for the first time that transplantation of UCMSC ameliorates ALI by enhancing the diminished levels of alveolar CD4(+)CD25(+) Foxp3(+)Treg and balancing anti- and pro-inflammatory factors in ALI mice.