Stem cells from foreign body granulation tissue accelerate recovery from acute kidney injury.

BACKGROUND: In previous studies, we obtained mesenchymal stem cells called granulation tissue stem cells (GTSC) from a regenerating granulation tissue created by placing a foreign body in the subcutaneous tissue of rats. Here, we used GTSC to ameliorate ischemia/reperfusion-induced acute kidney injury (AKI) in rats. METHODS: In two groups of Fischer rats, we induced ischemia/reperfusion injury. Group 1 (treated rats) received one intravenous injection of GTSC 3 h after injury; Group 2 (control rats) received vehicle. Both groups were subsequently studied by renal function tests, kidney histology and immunohistochemistry. RESULTS: At 24 and 48 h after injury, plasma creatinine and blood urea nitrogen were significantly lower in the treated rats as compared to control rats. The levels remained low and declined to near baseline levels by Day 4 in the treated group. At the cortico-medullary region, the treated rats showed significantly higher renal tubular cell proliferation and less tubular cell apoptosis. Histological analysis of the kidney for tubular dilatation, necrosis, congestion and casts was not significantly different in the two groups. To understand the mechanism of the GTSC effect, messenger RNA levels of several growth and immune modulatory factors were quantified in cultured GTSC and compared with those in cultured glomerular epithelial cell (GEC; a non-stem cell line). GTSC had 2- to 8-fold higher expression of FGF2, HGF, IGF-1, vascular endothelial growth factor (growth factors) and IL-4, IL-6 (anti-inflammatory factors) than GEC. CONCLUSIONS: Administration of GTSC accelerates recovery in rats with ischemia/reperfusion-induced AKI. This effect may be mediated by the paracrine action of growth and immune-suppressive factors secreted by these cells.

Omentum facilitates liver regeneration.

AIM: To investigate the mechanism of liver regeneration induced by fusing the omentum to a small traumatic injury created in the liver. We studied three groups of rats. In one group the rats were omentectomized; in another group the omentum was left in situ and was not activated, and in the third group the omentum was activated by polydextran particles. METHODS: We pre-activated the omentum by injecting polydextran particles and then made a small wedge wound in the rat liver to allow the omentum to fuse to the wound. We monitored the regeneration of the liver by determining the ratio of liver weight/body weight, by histological evaluation (including immune staining for cytokeratin-19, an oval cell marker), and by testing for developmental gene activation using reverse transcription polymerase chain reaction (RT-PCR). RESULTS: There was no liver regeneration in the omentectomized rats, nor was there significant regeneration when the omentum was not activated, even though in this instance the omentum had fused with the liver. In contrast, the liver in the rats with the activated omentum expanded to a size 50% greater than the original, and there was histologically an interlying tissue between the wounded liver and the activated omentum in which bile ducts, containing cytokeratin-19 positive oval cells, extended from the wound edge. In this interlying tissue, oval cells were abundant and appeared to proliferate to form new liver tissue. In rats pre-treated with drugs that inhibited hepatocyte growth, liver proliferation was ongoing, indicating that regeneration of the liver was the result of oval cell expansion. CONCLUSION: Activated omentum facilitates liver regeneration following injury by a mechanism that depends largely on oval cell proliferation.

Stromal cells cultured from omentum express pluripotent markers, produce high amounts of VEGF, and engraft to injured sites.

When rat omentum becomes activated by intraperitoneal injection of inert polydextran particles, these particles are rapidly surrounded by cells that express markers of adult stem cells (SDF-1alpha, CXCR4, WT-1) and of embryonic pluripotent cells (Oct-4, Nanog, SSEA-1). We have cultured such cells, because they may offer a convenient source of adult stem cells, and have found that they retain stem cell markers and produce high levels of vascular endothelial growth factor for up to ten passages. After systemic or local injection of these cultured cells into rats with acute injury of various organs, the cells specifically engraft at the injured sites. Thus, our experiments show that omental stromal cells can be cultured from activated omentum, and that these cells exhibit stem cell properties enabling them to be used for repair and possibly for the regeneration of damaged tissues.

Activated omentum becomes rich in factors that promote healing and tissue regeneration.

In order to study the mechanism by which an omental pedicle promotes healing when applied to an injured site, we injected a foreign body into the abdominal cavity to activate the omentum. One week after the injection, we isolated the omentum and measured blood vessel density, blood content, growth and angiogenesis factors (VEGF and others), chemotactic factors (SDF-1 alpha), and progenitor cells (CXCR-4, WT-1). We found that the native omentum, which consisted mostly of adipose tissue, expanded the mass of its non-adipose part (milky spots) 15- to 20-fold. VEGF and other growth factors increased by two- to four-fold, blood vessel density by three-fold, and blood content by two-fold. The activated omentum also showed increases in SDF-1 alpha, CXCR-4, and WT-1 cells (factors and cells positively associated with tissue regeneration). Thus, we propose that an omentum activated by a foreign body (or by injury) greatly expands its milky-spot tissue and becomes rich in growth factors and progenitor cells that facilitate the healing and regeneration of injured tissue.

Impaired integration of endothelial progenitor cells in capillaries of diabetic wounds is reversible with vascular endothelial growth factor infusion.

To understand impaired angiogenesis in diabetic wounds, polyvinyl tubes were implanted subcutaneously in rats to form a granulation tissue for 2 weeks and the granulation tissue was studied after inducing diabetes with streptozotocin. By 1 week of diabetes, the granulation tissue was bloody and thinner than controls, its medial layer was depleted of microvessels, and the surviving vessels appeared dehisced. Vascular endothelial growth factor (VEGF) in the diabetic granulation tissue was reduced by 50% compared with control granulation tissue. After 3 days of diabetes, the diabetic tissue showed a greater degree of apoptosis in the microvessels. Chemotactic factors [stromal cell-derived factor-1alpha and chemokine receptor-4 (CXCR-4)], responsible for attracting bone marrow cells, showed equal intensity in control and diabetic tissues. As expected, progenitor endothelial CD-34 cells were observed in abundance in both the control and the diabetic granulation tissues. However, although the CD-34-positive cells appeared mostly to be integrated in the blood vessels of the control tissue, fewer such cells were present in the blood vessels of the diabetic tissues, suggesting that CD-34 failed to integrate into new blood vessels. Infusion of VEGF in the granulation tissue of diabetic rats for 1 week resulted in complete prevention of the microvascular defect compared with the contralateral granulation tissue that showed the typical diabetic changes. It was concluded that diabetes causes reduction of VEGF in the wound, resulting in loss of blood vessels by apoptosis and possible failure of CD-34 cells to integrate into the vessel structure.

Transplanting fragments of diabetic pancreas into activated omentum gives rise to new insulin producing cells.

To determine if pancreatic progenitor cells can be induced to form insulin producing cells in vivo, we auto-transplanted fragments of streptozotocin-induced diabetic pancreas into omentum pre-injected with a foreign material. As shown previously, omentum pre-activated in this manner becomes rich in growth factors and progenitor cells. After auto-transplanting diabetic pancreas in the activated omentum, new insulin secreting cells appeared in the omentum in niches surrounding the foreign particles--a site previously shown to harbor progenitor cells. Extracts of these omenta contained measurable insulin. Four of eight diabetic animals treated in this manner became normoglycemic. This shows that new insulin producing cells can be regenerated from diabetic pancreas by auto-transplanting pancreatic fragments into the activated omentum, an environment rich in growth factors and progenitor cells.