Carlotta Reni

ABSTRACT We have previously shown that mice and human patients with diabetes mellitus develop microangiopathy in bone marrow (BM) resulting in depletion of stem cells (SCs). Since the endosteal niche was the most pauperised, we decided to investigate if activation of osteoclasts (OCLs) may participate in this process through detachment of SC from the endosteal niche leading to inappropriate mobilization. The study was performed in streptozotocin-induced type 1 diabetic (T1D) mice at 5, 11 and 20 weeks from induction of diabetes and in age-matched non-diabetic controls (C). Histochemical analysis of femoral bones at late stages of DM (20 weeks) showed a profound BM remodeling characterized by reduction of the hematopoietic component, fat accumulation and bone rarefaction. The activation of osteoclastogenesis was already seen at 5 weeks as assessed by counting the number of tartrate resistant acid phosphatase (TRAP) positive OCLs in the trabecular bone (2.9±0.3 vs. 1.6±0.7 cells/mm of endosteal length in C, p<0.01). Osteoclast activation was associated with a marked increase in peripheral blood lineage- Sca1+ cKit+ (LSK) cells (0.7±0.4 vs. 0.2±0.1% of total MNCs in C, p<0.05). Unexpectedly, mRNA and protein expression of Receptor Activator of NF-κB Ligand (RANKL), an inducer of OCL differentiation and activation, was reduced in total BM at 20 and 11 weeks (mRNA: 0.3±0.01 vs. 1.2±0.3 ddCT in C; Protein: 11.8±2 vs. 28.7±5 pg/ml, p<0.05 for both comparisons) thus implying that OCL activation occurs through a non-canonical mechanism. Interestingly, we found T1D induces acidosis in BM (pH: 6.9±0.02 vs. 7.0±0.03 in C, p<0.05). Moreover, similar pH changes in culturing media increased the number of BM-derived OCL-like cells in vitro (2.0±0.3 fold increase relative to C, p<0.05) suggesting a crucial role of acidosis in OCLs activation. In summary, parallel increases in circulating SCs and endosteal OCLs occur at initial phase of T1D, before any apparent alteration of hematopoiesis. OCL activation may lead to detachment of SCs from the endosteal niche and excessive mobilization, thus resulting in BM exhaustion at later stages. OCL-induced remodeling of the endosteal niche, which contains the most primitive SCs in BM, may contribute to the impairment of regenerative capacity in T1D.

Diabetes mellitus is considered a cardiovascular disease owing to its prevalent association with cardiovascular morbidity and mortality. Cardiovascular events are not only more frequent but also complicated with more severe outcomes in diabetic patients as compared with non-diabetic patients. One mechanism accounting for this difference consists of the impairment of the regenerative cellular machinery, which contributes to tissue healing. Recent evidence indicates the contribution of resident progenitor cells in post-ischemic tissue remodeling. In addition, a wide spectrum of cells from distant sources, including the bone marrow, is attracted and home to the healing tissue. Diabetes affects the process of mobilization and recruitment as well as intrinsic functional properties of bone marrow-derived progenitor cells. This review highlights current evidence for diabetes-induced damage of bone marrow hematopoietic progenitor cells in the endosteal and vascular niches.

Significance: Patients with diabetes mellitus suffer an excess of cardiovascular complications and recover worse from them as compared with their nondiabetic peers. It is well known that microangiopathy is the cause of renal damage, blindness, and heart attacks in patients with diabetes. This review highlights molecular deficits in stem cells and a supporting microenvironment, which can be traced back to oxidative stress and ultimately reduce stem cells therapeutic potential in diabetic patients. New research has shown that increased oxidative stress contributes to inducing microangiopathy in bone marrow (BM), the tissue contained inside the bones and the main source of stem cells. These precious cells not only replace old blood cells but also exert an important reparative function after acute injuries and heart attacks. The starvation of BM as a consequence of microangiopathy can lead to a less efficient healing in diabetic patients with ischemic complications. Furthermore, stem cells from a patient's BM are the most used in regenerative medicine trials to mend hearts damaged by heart attacks. A deeper understanding of redox signaling in BM stem cells will lead to new modalities for preserving local and systemic homeostasis and to more effective treatments of diabetic cardiovascular complications.

We investigated the association between the functional, epigenetic, and expressional profile of human adventitial progenitor cells (APCs) and therapeutic activity in a model of limb ischemia. Antigenic and functional features were analyzed throughout passaging in 15 saphenous vein (SV)-derived APC lines, of which 10 from SV leftovers of coronary artery bypass graft surgery and 5 from varicose SV removal. Moreover, 5 SV-APC lines were transplanted (8×10(5) cells, IM) in mice with limb ischemia. Blood flow and capillary and arteriole density were correlated with functional characteristics and DNA methylation/expressional markers of transplanted cells. We report successful expansion of tested lines, which reached the therapeutic target of 30 to 50 million cells in ≈10 weeks. Typical antigenic profile, viability, and migratory and proangiogenic activities were conserved through passaging, with low levels of replicative senescence. In vivo, SV-APC transplantation improved blood flow reco...

Vascular wall-resident progenitor cells hold great promise for cardiovascular regenerative therapy. This study evaluates the impact of oxidative stress on the viability and functionality of adventitia-derived progenitor cells (APCs) from vein remnants of coronary artery bypass graft (CABG) surgery. We also investigated the antioxidant enzymes implicated in the resistance of APCs to oxidative stress-induced damage and the effect of interfering with one of them, the extracellular superoxide dismutase (EC-SOD/SOD3), on APC therapeutic action in a model of peripheral ischemia. After exposure to hydrogen peroxide, APCs undergo apoptosis to a smaller extent than endothelial cells (ECs). This was attributed to up-regulation of antioxidant enzymes, especially SODs and catalase. Pharmacological inhibition of SODs increases reactive oxygen species (ROS) levels in APCs and impairs their survival. Likewise, APC differentiation results in SOD down-regulation and ROS-induced apoptosis. Oxidative stress increases APC migratory activity, while being inhibitory for ECs. In addition, oxidative stress does not impair APC capacity to promote angiogenesis in vitro. In a mouse limb ischemia model, an injection of naïve APCs, but not SOD3-silenced APCs, helps perfusion recovery and neovascularization, thus underlining the importance of this soluble isoform in protection from ischemia. This study newly demonstrates that APCs are endowed with enhanced detoxifier and antioxidant systems and that SOD3 plays an important role in their therapeutic activity in ischemia. APCs from vein remnants of CABG patients express antioxidant defense mechanisms, which enable them to resist stress. These properties highlight the potential of APCs in cardiovascular regenerative medicine.

Pain triggers a homeostatic alarm reaction to injury. It remains unknown, however, whether nociceptive signaling activated by ischemia is relevant for progenitor cells (PC) release from bone marrow. To this end, we investigated the role of the neuropeptide substance P (SP) and cognate neurokinin 1 (NK1) nociceptor in PC activation and angiogenesis during ischemia in mice and in human subjects. The mouse bone marrow contains sensory fibers and PC that express SP. Moreover, SP-induced migration provides enrichment for PC that express NK1 and promote reparative angiogenesis after transplantation in a mouse model of limb ischemia. Acute myocardial infarction and limb ischemia increase SP levels in peripheral blood, decrease SP levels in bone marrow, and stimulate the mobilization of NK1-expressing PC, with these effects being abrogated by systemic administration of the opioid receptor agonist morphine. Moreover, bone marrow reconstitution with NK1-knockout cells results in depressed PC ...

Vascular wall-resident progenitor cells hold great promise for cardiovascular regenerative therapy. This study evaluates the impact of oxidative stress on the viability and functionality of adventitia-derived progenitor cells (APCs) from vein remnants of coronary artery bypass graft (CABG) surgery. We also investigated the antioxidant enzymes implicated in the resistance of APCs to oxidative stress-induced damage and the effect of interfering with one of them, the extracellular superoxide dismutase (EC-SOD/SOD3), on APC therapeutic action in a model of peripheral ischemia. After exposure to hydrogen peroxide, APCs undergo apoptosis to a smaller extent than endothelial cells (ECs). This was attributed to up-regulation of antioxidant enzymes, especially SODs and catalase. Pharmacological inhibition of SODs increases reactive oxygen species (ROS) levels in APCs and impairs their survival. Likewise, APC differentiation results in SOD down-regulation and ROS-induced apoptosis. Oxidative stress increases APC migratory activity, while being inhibitory for ECs. In addition, oxidative stress does not impair APC capacity to promote angiogenesis in vitro. In a mouse limb ischemia model, an injection of naïve APCs, but not SOD3-silenced APCs, helps perfusion recovery and neovascularization, thus underlining the importance of this soluble isoform in protection from ischemia. This study newly demonstrates that APCs are endowed with enhanced detoxifier and antioxidant systems and that SOD3 plays an important role in their therapeutic activity in ischemia. APCs from vein remnants of CABG patients express antioxidant defense mechanisms, which enable them to resist stress. These properties highlight the potential of APCs in cardiovascular regenerative medicine.