**10. Bone marrow concentrates in critical limb ischemia**

Patients with significant, below the knee, vascular diseases and who are, first of all, not eligible for revascularization surgery or endovascular treatments due to several comorbidities or have high operative risk and had multiple failures of revascularization or high rate of re-stenosis, might be suitable candidates for biological cell-based therapy with BM-MSCs. Patients diagnosed with critical limb ischemia (CLI) might also suffer from chronic non-healing wounds, and the estimated amputation and mortality rates are high [138]. The application of regenerative medicine therapies, in particular the use of BM-MSCs protocols, has merged as a treatment option in patients with CLI. In these patients, the justification to use BMC is to promote the regeneration of impaired endothelium and stimulate neoangiogenesis in ischemic areas [139]. Several varieties of BM-MSC therapies have been studied in CLI patients, ranging from BM-derived mononuclear cells, CD34+ BM cells, to mesenchymal stromal cells. The outcomes of cell-based trials have been encouraging and demonstrated a significant decrease in the rate of amputation [140]. It can be concluded that BM-MSC applications have the potential to modify the natural history of intractable CLI, while high-quality randomized trials are needed [45].

### **11. Conclusions**

*Regenerative Medicine*

**8.1 Degenerative disk disease**

and behavioral therapy.

option to augment PLF healing.

**9. Bone marrow concentrates in chronic wounds**

**8.2 Spinal surgery**

pain is chronic. Nowadays, autologous regenerative applications have been made

Pettine et al. studied the use of intra-discal BMC injections in patients with DDD [131]. The authors injected 26 symptomatic patients for lumbar diskogenic back pain and disability and evaluated their postinjection outcomes using disability scores, pain scores, and MRIs. At 2-year follow-up, patients experienced significant improvements in disability and pain scores. This group was the first to report on MSC dose-dependent outcome responses. Patients receiving greater concentrations of autologous BM-MSC (expressed as CFU-Fs > 2000/ml) experienced a faster and greater reduction in pain scores. Later, these findings were strengthened with a follow-up study at 36 months, showing similar outcome results [43]. At 5-year follow-up, absolute and percentage reductions in pain and disability scores were sustained, with no adverse events reported through the 5-year follow-up period [132]. The American Society of Interventional Pain Physicians published recently guidelines addressing responsible and safe use of autologous biologics in the management of lower back pain [133]. Their extensive analysis revealed that there is level III evidence for the use of PRP and BM-MSCs. The guidelines also state that, following diagnostic evidence, regenerative therapies should be provided to patients as an independent therapy. If appropriate and indicated, regenerative therapies can be combined with conventional medicinal therapy or in conjunction with physical

Hart et al. informed on a prospective, randomized, and blinded study in patients with lumbar disease the use of BMC mixed with allograft spongiosa chips during surgical posterolateral fusion (PLF) procedures. Patients underwent instrumented lumbar spine PLF procedures [134]. Fusion status and the degree of mineralization were evaluated by two radiologists blinded to patient group affiliation. X-ray examination, in control patients at 12-month follow-up, showed that the bone graft mass fused in none of the cases and, at 24-months, in four cases (10%). In the BMC treatment group, 6 cases (15%) achieved fusion at 12 months and 14 cases (35%) at 24 months. Computed tomography scans showed that 40% of control patients and 80% of BMC-treated patients had evidence of at least a unilateral continuous bridging of the bones between neighboring vertebrae at 24 months, significantly favoring the mixture of spongiosa bone with autologous BMC (*P* < 0.05) as an efficient

Cell-based therapies are an attractive approach for the treatment of recalcitrant chronic wounds. BM-MSCs have been studied as a therapeutic strategy in chronic hard-to-heal wounds [135]. The orchestrated process of wound healing entails cellular and hormonal physiological processes in inflammation, proliferation, collagen matrix formation, and epithelialization which are regulated by various platelet-derived growth factors, such as TGF-b, VEGF, PDGF, granulocyte-

macrophage colony-stimulating factor, the interleukin family, EGF, FGF, and TNF-a [44, 105]. In chronic, poor-healing wounds, the activity and effectivity of growth factors and cytokines are often reduced due to a chronically inflamed wound.

available to patients as an alternative treatment option.

**28**

Regenerative medicine technologies offer solutions to a number of compelling clinical problems that have not been able to adequately result in a solution through the use of drugs, surgery, or permanent replacement devices. Reviewing the last decades regarding autologous biological therapies, BM-MSCs have gained great interest. The purpose of this chapter was to review specific characteristics of bone marrow tissue and its cellular content, in particular the mesenchymal stem cells. Considerations when performing aspiration techniques and bone marrow concentrate preparations were presented, including explicit roles of hematopoietic and mesenchymal stem cells and other cytokines. Among autologous tissue-based cellular therapies, bone marrow mesenchymal cell therapies have been the most frequently employed and reported on, despite the fact that effects of coadjuvants, dosing, repetitive procedures, etc. are not yet established. Cultured MSC therapeutic interventions require strict procedures and biological license agreements, making them less attractive for same-day regenerative therapies. Using at POC BM-MSC concentrates overcomes these lengthy regulatory processes without the need for mandates. Clinical translation of BM-MSC-based therapies remains a work in progress, as proper standardization has not yet been recognized [53]. However, in the clinical setting, effective and safe autologous BMA harvesting and preparation of BMC have been reported [42]. More, better, organized randomized clinical trials that are warranted with accurate follow-up data revealing the efficacy of BM-MSC therapy, including laboratory validation of the used products, should be a future goal. Furthermore, proper deliberations should manage the enormous variability aspects, like aspiration techniques, imaging options and procedures, BMC preparation protocols, effect of patient age, as well as tissue disease state. Therapy failures should also be highlighted in order to understand how they impact the therapy outcomes. Ultimately, the adoption of an accepted standard of overall regenerative biological preparations, including critical and ambivalent nuances, is crucial for future regenerative medicine practices.
