Vladimir Serikov

Intrapulmonary Delivery of Bone Marrow-Derived Mesenchymal Stem Cells Improves Survival and Attenuates Endotoxin-Induced Acute Lung Injury in Mice

Recent in vivo and in vitro work suggests that mesenchymal stem cells (MSC) have anti-inflammatory properties. In this study, we tested the effect of administering MSC directly into the airspaces of the lung 4 h after the intrapulmonary administration of Escherichia coli endotoxin (5 mg/kg). MSC increased survival compared with PBS-treated control mice at 48 h (80 vs 42%; p < 0.01). There was also a significant decrease in excess lung water, a measure of pulmonary edema (145 ± 50 vs 87 ± 20 μl; p < 0.01), and bronchoalveolar lavage protein, a measure of endothelial and alveolar epithelial permeability (3.1 ± 0.4 vs 2.2 ± 0.8 mg/ml; p < 0.01), in the MSC-treated mice. These protective effects were not replicated by the use of further controls including fibroblasts and apoptotic MSC. The beneficial effect of MSC was independent of the ability of the cells to engraft in the lung and was not related to clearance of the endotoxin by the MSC. MSC administration mediated a down-regulation of proinflammatory responses to endotoxin (reducing TNF-α and MIP-2 in the bronchoalveolar lavage and plasma) while increasing the anti-inflammatory cytokine IL-10. In vitro coculture studies of MSC with alveolar macrophages provided evidence that the anti-inflammatory effect was paracrine and was not cell contact dependent. In conclusion, treatment with intrapulmonary MSC markedly decreases the severity of endotoxin-induced acute lung injury and improves survival in mice.

Allogeneic human mesenchymal stem cells for treatment of E. coli endotoxin-induced acute lung injury in the ex vivo perfused human lung

Recent studies have suggested that bone marrow-derived multipotent mesenchymal stem cells (MSCs) may have therapeutic applications in multiple clinical disorders including myocardial infarction, diabetes, sepsis, and hepatic and acute renal failure. Here, we tested the therapeutic capacity of human MSCs to restore alveolar epithelial fluid transport and lung fluid balance from acute lung injury (ALI) in an ex vivo perfused human lung preparation injured by E. coli endotoxin. Intra-bronchial instillation of endotoxin into the distal airspaces resulted in pulmonary edema with the loss of alveolar epithelial fluid transport measured as alveolar fluid clearance. Treatment with allogeneic human MSCs or its conditioned medium given 1 h following endotoxin-induced lung injury reduced extravascular lung water, improved lung endothelial barrier permeability and restored alveolar fluid clearance. Using siRNA knockdown of potential paracrine soluble factors, secretion of keratinocyte growth factor was essential for the beneficial effect of MSCs on alveolar epithelial fluid transport, in part by restoring amiloride-dependent sodium transport. In summary, treatment with allogeneic human MSCs or the conditioned medium restores normal fluid balance in an ex vivo perfused human lung injured by E. coli endotoxin.

Antibacterial Effect of Human Mesenchymal Stem Cells Is Mediated in Part from Secretion of the Antimicrobial Peptide LL-37

Recent in vivo studies indicate that mesenchymal stem cells (MSCs) may have beneficial effects in the treatment of sepsis induced by bacterial infection. Administration of MSCs in these studies improved survival and enhanced bacterial clearance. The primary objective of this study was to test the hypothesis that human MSCs possessed intrinsic antimicrobial properties. We studied the effect of human MSCs derived from bone marrow on the bacterial growth of Gram‐negative (Escherichia coli and Pseudomonas aeruginosa) and Gram‐positive (Staphylococcus aureus) bacteria. MSCs as well as their conditioned medium (CM) demonstrated marked inhibition of bacterial growth in comparison with control medium or normal human lung fibroblasts (NHLF). Analysis of expression of major antimicrobial peptides indicated that one of the factors responsible for the antimicrobial activity of MSC CM against Gram‐negative bacteria was the human cathelicidin antimicrobial peptide, hCAP‐18/LL‐37. Both m‐RNA and protein expression data showed that the expression of LL‐37 in MSCs increased after bacterial challenge. Using an in vivo mouse model of E. coli pneumonia, intratracheal administration of MSCs reduced bacterial growth (in colony‐forming unit) in the lung homogenates and in the bronchoalveolar lavage (BAL) fluid, and administration of MSCs simultaneously with a neutralizing antibody to LL‐37 resulted in a decrease in bacterial clearance. In addition, the BAL itself from MSC‐treated mice had a greater antimicrobial activity in comparison with the BAL of phosphate buffered saline (PBS)‐treated mice. Human bone marrow‐derived MSCs possess direct antimicrobial activity, which is mediated in part by the secretion of human cathelicidin hCAP‐18/ LL‐37. STEM CELLS 2010;28:2229–2238

Mesenchymal stem cells enhance survival and bacterial clearance in murine Escherichia coli pneumonia

Rationale Bacterial pneumonia is the most common infectious cause of death worldwide and treatment is increasingly hampered by antibiotic resistance. Mesenchymal stem cells (MSCs) have been demonstrated to provide protection against acute inflammatory lung injury; however, their potential therapeutic role in the setting of bacterial pneumonia has not been well studied. Objective This study focused on testing the therapeutic and mechanistic effects of MSCs in a mouse model of Gram-negative pneumonia. Methods and results Syngeneic MSCs from wild-type mice were isolated and administered via the intratracheal route to mice 4 h after the mice were infected with Escherichia coli. 3T3 fibroblasts and phosphate-buffered saline (PBS) were used as controls for all in vivo experiments. Survival, lung injury, bacterial counts and indices of inflammation were measured in each treatment group. Treatment with wild-type MSCs improved 48 h survival (MSC, 55%; 3T3, 8%; PBS, 0%; p<0.05 for MSC vs 3T3 and PBS groups) and lung injury compared with control mice. In addition, wild-type MSCs enhanced bacterial clearance from the alveolar space as early as 4 h after administration, an effect that was not observed with the other treatment groups. The antibacterial effect with MSCs was due, in part, to their upregulation of the antibacterial protein lipocalin 2. Conclusions Treatment with MSCs enhanced survival and bacterial clearance in a mouse model of Gram-negative pneumonia. The bacterial clearance effect was due, in part, to the upregulation of lipocalin 2 production by MSCs.

Potential application of mesenchymal stem cells in acute lung injury.

Despite extensive research into the pathogenesis of acute lung injury and the acute respiratory distress syndrome (ALI/ARDS), mortality remains high at approximately 40%. Current treatment is primarily supportive, with lung-protective ventilation and a fluid conservative strategy. Pharmacologic therapies that reduce the severity of lung injury in experimental studies have not yet been translated into effective clinical treatment options. Therefore, innovative therapies are needed. Recent studies have suggested that bone-marrow-derived multipotent mesenchymal stem cells (MSC) may have therapeutic applications in multiple clinical disorders including myocardial infarction, diabetes, sepsis, hepatic and acute renal failure. Recently, MSC have been studied in several in vivo models of lung disease. This review focuses on first describing the existing experimental literature that has tested the use of MSC in models of ALI/ARDS, and then the potential mechanisms underlying their therapeutic use with an emphasis on secreted paracrine soluble factors. The review concludes with a discussion of future research directions required for potential clinical trials.

Human term placenta as a source of hematopoietic cells.

The main barrier to a broader clinical application of umbilical cord blood (UCB) transplantation is its limiting cellular content. Thus, the discovery of hematopoietic progenitor cells in murine placental tissue led us investigate whether the human placenta contains hematopoietic cells, sites of hematopoiesis, and to develop a procedure of processing and storing placental hematopoietic cells for transplantation. Here we show that the human placenta contains large numbers of CD34-expressing hematopoietic cells, with the potential to provide a cellular yield several-fold greater than that of a typical UCB harvest. Cells from fresh or cryopreserved placental tissue generated erythroid and myeloid colonies in culture, and also produced lymphoid cells after transplantation in immunodeficient mice. These results suggest that human placenta could become an important new source of hematopoietic cells for allogeneic transplantation.The main barrier to a broader clinical application of umbilical cord blood (UCB) transplantation is its limiting cellular content. Thus, the discovery of hematopoietic progenitor cells in murine placental tissue led us investigate whether the human placenta contains hematopoietic cells, sites of hematopoiesis, and to develop a procedure of processing and storing placental hematopoietic cells for transplantation. Here we show that the human placenta contains large numbers of CD34-expressing hematopoietic cells, with the potential to provide a cellular yield several-fold greater than that of a typical UCB harvest. Cells from fresh or cryopreserved placental tissue generated erythroid and myeloid colonies in culture, and also produced lymphoid cells after transplantation in immunodeficient mice. These results suggest that human placenta could become an important new source of hematopoietic cells for allogeneic transplantation.

Multipotent Stromal Stem Cells from Human Placenta Demonstrate High Therapeutic Potential

We describe human chorionic mesenchymal stem cell (hCMSC) lines obtained from the chorion of human term placenta with high therapeutic potential in human organ pathology. hCMSCs propagated for more than 100 doublings without a decrease in telomere length and with no telomerase activity. Cells were highly positive for the embryonic stem cell markers OCT‐4, NANOG, SSEA‐3, and TRA‐1–60. In vitro, cells could be differentiated into neuron‐like cells (ectoderm), adipocytes, osteoblasts, endothelial‐like cells (mesoderm), and hepatocytes (endoderm)—derivatives of all three germ layers. hCMSCs effectively facilitated repair of injured epithelium as demonstrated in an ex vivo‐perfused human lung preparation injured by Escherichia coli endotoxin and in in vitro human lung epithelial cultures. We conclude that the chorion of human term placenta is an abundant source of multipotent stem cells that are promising candidates for cell‐based therapies.

A BRIEF COMMUNICATION

The main barrier to a broader clinical application of umbilical cord blood (UCB) transplantation is its limiting cellular content. Thus, the discovery of hematopoietic progenitor cells in murine placental tissue led us investigate whether the human placenta contains hematopoietic cells, sites of hematopoiesis, and to develop a procedure of processing and storing placental hematopoietic cells for transplantation. Here we show that the human placenta contains large numbers of CD34-expressing hematopoietic cells, with the potential to provide a cellular yield several-fold greater than that of a typical UCB harvest. Cells from fresh or cryopreserved placental tissue generated erythroid and myeloid colonies in culture, and also produced lymphoid cells after transplantation in immunodeficient mice. These results suggest that human placenta could become an important new source of hematopoietic cells for allogeneic transplantation.The main barrier to a broader clinical application of umbilical cord blood (UCB) transplantation is its limiting cellular content. Thus, the discovery of hematopoietic progenitor cells in murine placental tissue led us investigate whether the human placenta contains hematopoietic cells, sites of hematopoiesis, and to develop a procedure of processing and storing placental hematopoietic cells for transplantation. Here we show that the human placenta contains large numbers of CD34-expressing hematopoietic cells, with the potential to provide a cellular yield several-fold greater than that of a typical UCB harvest. Cells from fresh or cryopreserved placental tissue generated erythroid and myeloid colonies in culture, and also produced lymphoid cells after transplantation in immunodeficient mice. These results suggest that human placenta could become an important new source of hematopoietic cells for allogeneic transplantation.

Evidence of Temporary Airway Epithelial Repopulation and Rare Clonal Formation by BM‐derived Cells Following Naphthalene Injury in Mice

The goal of the study was to investigate participation of bone marrow (BM) cells in the process of airway epithelial restoration after naphthalene‐induced injury. We transplanted sex‐mismatched green fluorescent protein (GFP) ‐tagged BM‐derived cultured plastic‐adherent mesenchymal stem cells into 5Gy‐irradiated C57BL/6 recipients. After 1 month of recovery, experimental animals were subjected to 250 mg/kg naphthalene IP. Animals were killed at 2–30 days after naphthalene. By immunofluorescence, immunohistochemistry, and by in situ hybridization for the Y‐chromosome, we observed patches of donor‐derived cells in the large and small conducting airways, mostly at 2–6 days after injury. GFP+ cells in the epithelium of airways were positive for pancytokeratin and some other epithelial markers. Although rare, GFP+ cells formed clear isolated patches of the bronchial epithelium, consistent with clonal formation; as some cells were also positive for proliferating cell nuclear antigen, a marker of proliferating cells. After day 12, only occasional GFP+ cells were present in the epithelium. These data confirm that bone marrow‐derived cultured mesenchymal cells can participate in the recovery of the injured airway epithelium after naphthalene‐induced injury with minimal long‐term engraftment. Anat Rec, 2007.

Constitutive expression of the human peroxiredoxin V gene contributes to protection of the genome from oxidative DNA lesions and to suppression of transcription of noncoding DNA

Peroxiredoxins belong to a family of antioxidant proteins that neutralize reactive oxygen species. One member of this family, peroxiredoxin I (PRDX1), suppresses DNA oxidation. Peroxiredoxin V (PRDX5) has been cloned as a transcriptional corepressor, as a peroxisomal/mitochondrial antioxidant protein, and as an inhibitor of p53‐dependent apoptosis. Promoters of mammalian PRDX5 genes contain clusters of antioxidant response elements, which can bind the transcription factor NRF2. However, we found that expression of the human PRDX5 gene in situ was not stimulated by the oxidative agent menadione. Silencing of the NRF2 gene in the absence of oxidative stress by specific siRNA did not decrease PRDX5 protein concentration. We also constructed clones of human lung epithelial cells A549 with siRNA‐mediated knockdown of the PRDX5 gene. This led to a significant increase in 8‐oxoguanine formation in cell DNA. In the PRDX5 knockdown clone, an increase in transcripts containing sequences of alpha‐satellite and satellite III DNAs was also detected, suggesting that this protein may be required for silencing of heterochromatin. Together, these results suggest that constitutively expressed PRDX5 gene plays an important role in protecting the genome against oxidation and may also be involved in the control of transcription of noncoding DNA.