Youngsook Son

A new role of substance P as an injury-inducible messenger for mobilization of CD29 + stromal-like cells

Tissue injury may create a specific microenvironment for inducing the systemic participation of stromal-like cells in the repair process. Here we show that substance P is an injury-inducible factor that acts early in the wound healing process to induce CD29+ stromal-like cell mobilization. Likewise, mobilization of such cells also occurs in uninjured mice, rats and rabbits if substance P is intravenously injected. Upon further characterization these substance P–mobilized CD29+ cells were found to be similar to stromal cells from a number of connective tissues, including bone marrow (that is, bone marrow stromal cells, or BMSCs). Both substance P injection and transfusion of autologously derived substance P–mobilized CD29+ cells from uninjured rabbits accelerated wound healing in an alkali burn model. Also, epithelial engraftment of the transfused cells into the injured tissue occurred during the wound healing. Finally, using human BMSCs as a test population, we show that substance P stimulates transmigration, cell proliferation, activation of the extracellular signal–related kinases (Erk) 1 and 2 and nuclear translocation of β-catenin in vitro. This finding highlights a previously undescribed function of substance P as a systemically acting messenger of injury and a mobilizer of CD29+ stromal-like cells to participate in wound healing.

DNA damage in the kidneys of diabetic rats exhibiting microalbuminuria

8-Hydroxydeoxyguanosine (8-OHdG), an oxygen radical induced modification of purine residue in DNA, was measured in the liver, pancreas, and kidney of streptozotocin-induced diabetic rats (STZR) exhibiting microalbuminuria. At 4 weeks after the injection of streptozotocin (50 mg/kg, i.v.), the rate of urinary albumin excretion was 0.5 +/- 0.1 and 2.0 +/- 0.2 mg/24 h in age-matched control rats (CR) and STZR, respectively. Compared to CR, STZR also showed a significantly increased level of 8-OHdG in the kidney but not the liver and pancreas. Amounts of 8-OHdG/10(5) dG for CR and STZR were 3.4 +/- 0.3 and 5.1 +/- 0.2 for renal cortices, and 4.1 +/- 0.2 and 20.0 +/- 3.7 for renal papillae. Daily injection of insulin (2 U, SC) starting on the third day after streptozotocin treatment significantly reduced both urinary albumin excretion and papillary 8-OHdG formation, which suggests that these are associated with the diabetic state induced by streptozotocin rather than a direct nephrotoxic effect of the drug. This study suggests that formation of 8-OHdG and, therefore, oxidative damage are closely related in the process of diabetic nephropathy.

Substance P induces M2-type macrophages after spinal cord injury.

The potential benefits or the tissue-damaging effects of inflammatory response after central nervous system injuries have long been disputed. Recent studies have noted that substance P (SP), a neuropeptide, plays an important role in the wound-healing process by recruiting bone marrow stem cells to the injured tissue. In this study, we examined whether SP can enhance recovery from spinal cord injury (SCI) in Sprague–Dawley rats through its known function of stem cell mobilization and/or through the modulation of inflammation. We examined proinflammatory and anti-inflammatory cytokines and markers for macrophage subtypes. SP treatment modulated the SCI microenvironment toward a more anti-inflammatory and reparative one by inducing interleukin-10 and M2 macrophages and suppressing inducible nitric oxide synthase and tumor necrosis factor-&agr;. This modulation was achieved at 1 day much earlier than SP-stimulated bone marrow stem cells’ mobilization. Early intervention of the devastating inflammatory response by SP treatment caused the lesion cavity to become filled with robust axonal outgrowth that overlaid the M2 macrophages at 2 weeks – all of which culminated in tissue sparing and improvement in functional recovery from the SCI. SP is therefore a potential anti-inflammatory modulator for the treatment of injury-induced inflammatory central nervous system disorders.

Schwann cells differentiated from spheroid-forming cells of rat subcutaneous fat tissue myelinate axons in the spinal cord injury

In the present study, we found that nestin-expressing spheroid cells derived from multipotent adipose stem cells of subcutaneous fat tissue could efficiently differentiate into Schwann cells (SCs) in vitro based on expression of SC markers such as A2B5, GFAP, O4, p75, S100, Sox10, Krox-20, and L1. The induced SC is engrafted to spinal cord injury lesions and formed a peripheral nervous system (PNS)-type myelin sheath on central nervous system (CNS) axons. PNS-type myelin sheath formation in repaired tissue was confirmed by transplantation of both induced PKH26-labeled SC and induced EGFP-expressing SC generated from EGFP transgenic rats. In addition to direct participation as myelin sheath-forming SC in repaired tissue, the induced SC also expressed several neurotrophic factors, as did native SC, which may suggest an additional role for induced SC in stimulation of endogenous healing responses. Thus, spheroid-forming cells from subcutaneous fat tissue demonstrated rapid and efficient induction into SC, and such cells show therapeutic promise for repair of damage to the CNS and PNS.

Perspectives on mesenchymal stem cells: Tissue repair, immune modulation, and tumor homing

Mesenchymal stem cells (MSCs) or MSC-like cells have been identified in a variety of different tissues that share molecular expression profiles and biological functions but also retain a unique differentiation preference depending on their tissue origins. MSCs play beneficial roles in the healing of damaged tissue by directly differentiating to many different resident cell types and/or by secreting several trophic factors that aid tissue repair. Aside from MSCs’ reparative stem cell function, they drive immune responses toward immunosuppression and anti-inflammation. This novel function of MSCs opens up new avenues for clinical development of MSC immune-therapeutics to treat uncontrollable, life threatening, severe, chronic inflammation and autoimmune disease. Unexpectedly high rates of MSCs’ tumor homing capacity and their tumor supporting capability have also been noted in tumor-bearing animal models. In this review, we will discuss MSCs’ basic cell biology and perspectives on MSCs in terms of tissue repair, immune modulation, and tumor homing.

Ocular Surface Reconstruction With Autologous Nasal Mucosa in Cicatricial Ocular Surface Disease

PURPOSE To investigate the possibility of replacing the metaplastic ocular surface with nasal mucosa, and to evaluate the results of autologous nasal and oral mucosal transplantation in cicatricial ocular surface diseases. DESIGN Retrospective interventional case series. METHODS We studied 6 eyes in 6 patients with chemical burns, which were characterized by a cicatricial ocular surface. After removal of cicatricial tissues and symblepharolysis, autologous nasal mucosa was transplanted in all patients. In 3 patients with extensive damage, oral mucosal autografting was performed concurrently. The nasal and oral mucosa was evaluated using immunohistochemical analysis for p63, K3, MUC5AC, and CD34. Clinical outcomes were assessed based on visual acuity, ocular manifestations, and liquid-based cytology. RESULTS Immunohistochemical analysis revealed a plentitude of p63 and K3 in nasal mucosal epithelium. Goblet cells and MUC5AC expression were only observed in nasal mucosal epithelium, not in oral mucosal epithelium. Well-developed parallel vasculature was demonstrated in the nasal mucosa. In contrast, perpendicular vasculature was demonstrated in the oral mucosa. This vascular feature remained after transplantations. In all patients, ocular surface stability recovered with no major complications and increased goblet cells were observed on ocular surface. However, delayed epithelialization and ischemic thinning were seen at oral mucosal graft sites. CONCLUSIONS Nasal mucosa, which has the advantage of well-developed parallel vasculature, enriched goblet cells, and plenty of stem cells, may be an ideal substitute for a cicatricial ocular surface. Transplantation of autologous nasal mucosa is a very effective method for achieving ocular surface reconstruction in cicatricial ocular surface diseases.

Substance P stimulates the recovery of bone marrow after the irradiation.

The therapeutic use of ionizing radiation (e.g., X‐rays and γ‐rays) needs to inflict minimal damage on non‐target tissue. Recent studies have shown that substance P (SP) mediates multiple activities in various cell types, including cell proliferation, anti‐apoptotic responses, and inflammatory processes. The present study investigated the effects of SP on γ‐irradiated bone marrow stem cells (BMSCs). In mouse bone marrow extracts, SP prolonged activation of Erk1/2 and enhanced Bcl‐2 expression, but attenuated the activation of apoptotic molecules (e.g., p38 and cleaved caspase‐3) and down‐regulated Bax. We also observed that SP‐decreased apoptotic cell death and stimulated cell proliferation in γ‐irradiated mouse bone marrow tissues through TUNEL assay and PCNA analysis. To determine how SP affects bone marrow stem cell populations, mouse bone marrow cells were isolated and colony‐forming unit (CFU) of mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) was estimated. SP‐pretreated ones showed higher CFUs of MSC and HSC than untreated ones. Furthermore, when SP was pretreated in cultured human MSC, it significantly decreased apoptotic cells at 48 and 72 h after γ‐irradiation. Compared with untreated cells, SP‐treated human MSCs showed reduced cleavage of apoptotic molecules such as caspase‐8, ‐9, ‐3, and poly ADP‐ribose polymerase (PARP). Thus, our results suggest that SP alleviates γ‐radiation‐induced damage to mouse BMSCs and human MSCs via regulation of the apoptotic pathway. J. Cell. Physiol. 226: 1204–1213, 2011.

A new 3D reconstituted human corneal epithelium model as an alternative method for the eye irritation test.

Many efforts are being made to develop new alternative in vitro test methods for the eye irritation test. Here we report a new reconstructed human corneal epithelial model (MCTT HCE model) prepared from primary-cultured human limbal epithelial cells as a new alternative in vitro eye irritation test method. In histological and immunohistochemical observation, MCTT HCE model displayed a morphology and biomarker expressions similar to intact human cornea. Moreover, the barrier function was well preserved as measured by high transepithelial electrical resistance, effective time-50 for Triton X-100, and corneal thickness. To employ the model as a new alternative method for eye irritation test, protocol refinement was performed and optimum assay condition was determined including treatment time, treatment volume, post-incubation time and rinsing method. Using the refined protocol, 25 reference chemicals with known eye irritation potentials were tested. With the viability cut-off value at 50%, chemicals were classified to irritant or non-irritant. When compared with GHS classification, the MCTT HCE model showed the accuracy of 88%, sensitivity of 100% and specificity of 77%. These results suggest that the MCTT HCE model might be useful as a new alternative eye irritation test method.

Substance P ameliorates collagen II-induced arthritis in mice via suppression of the inflammatory response.

Current rheumatoid arthritis (RA) therapies such as biologics inhibiting pathogenic cytokines substantially delay RA progression. However, patient responses to these agents are not always complete and long lasting. This study explored whether substance P (SP), an 11 amino acids long endogenous neuropeptide with the novel ability to mobilize mesenchymal stem cells (MSC) and modulate injury-mediated inflammation, can inhibit RA progression. SP efficacy was evaluated by paw swelling, clinical arthritis scoring, radiological analysis, histological analysis of cartilage destruction, and blood levels of tumor necrosis factor-alpha (TNF-α) interleukin (IL)-10, and IL-17 in vivo. SP treatment significantly reduced local inflammatory signs, mean arthritis scores, degradation of joint cartilage, and invasion of inflammatory cells into the synovial tissues. Moreover, the SP treatment markedly reduced the size of spleens enlarged by excessive inflammation in CIA, increased IL-10 levels, and decreased TNF-α and IL-17 levels. Mobilization of stem cells and induction of T(reg) and M2 type macrophages in the circulation were also increased by the SP treatment. These effect of SP might be associated with the suppression of inflammatory responses in RA and, furthermore, blockade of RA progression. Our results propose SP as a potential therapeutic for autoimmune-related inflammatory diseases.

A new paradigm for stem cell therapy: Substance-P as a stem cell-stimulating agent

Bone marrow is a reservoir for hematopoietic stem cells, endothelial precursor cells, and bone marrow stromal cells (also generally called mesenchymal stem cells), whose positive role in tissue repair is highly anticipated. In this report, we introduce a novel function of substance-P (SP), an 11-amino-acid peptide, as an injury-inducible messenger to mobilize bone marrow stem cells to the blood and finally to engage in tissue repair. This new drug may substitute for ex vivo cell culture of therapeutic cells by stimulating cell proliferation in the bone marrow in vivo and mobilizing those therapeutic cells to the patient’s own blood stream. Again, the additional role of SP in mitigating inflammation-mediated tissue damage can further rationalize the clinical development of SPpeptide as a stem cell stimulant.