Angiogenic effects of cell therapy within a biomaterial scaffold in a rat hind limb ischemia model.

Critical limb ischemia (CLI) is a life- and limb-threatening condition affecting 1-10% of humans worldwide with peripheral arterial disease. Cellular therapies, such as bone marrow-derived mesenchymal stem cells (MSCs) have been used for the treatment of CLI. However, little information is available regarding the angiogenic potency of MSCs and mast cells (MC) in angiogenesis. The aim of this study was to evaluate the ability of MCs and MSCs to induce angiogenesis in a rat model of ischemic hind limb injury on a background of a tissue engineered hydrogel scaffold. Thirty rats were randomly divided into six control and experimental groups as follows: (a) Control healthy (b) Ischemic positive control with right femoral artery transection, (c) ischemia with hydrogel scaffold, (d) ischemia with hydrogel plus MSC, (e) ischemia with hydrogel plus MC and (f) ischemia with hydrogel plus MSC and MCs. 106 of each cell type, isolated from bone marrow stroma, was injected into the transected artery used to induce hind limb ischemia. The other hind limb served as a non-ischemic control. After 14 days, capillary density, vascular diameter, histomorphometry and immunohistochemistry at the transected location and in gastrocnemius muscles were evaluated. Capillary density and number of blood vessels in the region of the femoral artery transection in animals receiving MSCs and MCs was increased compared to control groups (P < 0.05). Generally the effect of MCs and MSCs was similar although the combined MC/MSC therapy resulted in a reduced, rather than enhanced, effect. In the gastrocnemius muscle, immunohistochemical and histomorphometric observation showed a great ratio of capillaries to muscle fibers in all the cell-receiving groups (P < 0.05). The data indicates that the combination of hydrogel and cell therapy generates a greater angiogenic potential at the ischemic site than cell therapy or hydrogels alone.

The Role of Mast Cells in IgE-Independent Lung Diseases.

Mast cells (MCs) are granular cells of the innate immune system which develop from CD34+/CD117+ progenitors and play a role in orchestrating adaptive immune responses. They have a well-known role in allergic reactions following immunoglobulin (Ig)E-mediated activation of the cell-surface expressed IgE high-affinity receptor (FcεRI). MCs can also respond to various other stimuli due to the expression of a variety of receptors including toll-like receptors (TLRs), immunoglobulin (IgG) receptors (FcγR), complement receptors such as C5a (CD88) expressed by skin MCs, neuropeptides receptors including nerve growth factor receptor, (NGFR), cytokines receptors such as (IL)-1R and IL-3R, and chemokines receptors including CCR-1 and CCR-3. MCs release three groups of mediators upon degranulation differentiated according to their chemical composition, storage, and time to release. These include preformed mediators (mainly histamine, tryptase, and chymase), de novo synthesized mediators such as prostaglandin (PG)D2, leukotriene (LT)B4 and LTD4, and cytokines including IL-1ß, IL-3, tumor necrosis factor (TNF)α, and transforming growth factor(TGF)-ß. Emerging evidence indicates a role for IgE-independent MC activation in the late-stage asthmatic response as well as in non-allergic airway diseases including chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF), and lung cancer. MC infiltration/activation has been reported in some, but not all, studies of lung cancer. MC-derived TNF-α possesses tumor-suppressive activity while IL-1ß supports tumor progression and metastasis. In IPF lungs, an increase in density of tryptase- and chymase-positive MCs (MCTC) and overexpression of TGF-ß support the fibrosis progression. MC-derived chymase activates latent TGF-ß that induces the differentiation of fibroblasts to matrix-producing myofibroblasts. In summary, increasing evidence highlights a critical role of MCs in non-allergic diseases that may indicate new approaches for therapy.

Histological Evidence for Therapeutic Induction of Angiogenesis Using Mast Cells and Platelet-Rich Plasma within A Bioengineered Scaffold following Rat Hindlimb Ischemia.

OBJECTIVE: Peripheral arterial disease results from obstructed blood flow in arteries and increases the risk of amputation in acute cases. Therapeutic angiogenesis using bioengineered tissues composed of a chitosan scaffold that was enriched with mast cells (MCs) and/or platelet-rich plasma (PRP) was used to assess the formation of vascular networks and subsequently improved the functional recovery following hindlimb ischemia. This study aimed to find an optimal approach for restoring local vascularization. MATERIALS AND METHODS: In this experimental study, thirty rats were randomly divided into six experimental groups: a. Ischemic control group with right femoral artery transection, b. Ischemia with phosphate-buffered saline (PBS) control group, c. Ischemia with chitosan scaffold, d. Ischemia with chitosan and MCs, e. Ischemia with chitosan and PRP, and f. Ischemia with chitosan, PRP, and MCs. The left hind limbs served as non-ischemic controls. The analysis of capillary density, arterial diameter, histomorphometric analysis and immunohistochemistry at the transected locations and in gastrocnemius muscles was performed. RESULTS: The group treated with chitosan/MC significantly increased capillary density and the mean number of large blood vessels at the site of femoral artery transection compared with other experimental groups (P<0.05). The treatment with chitosan/MC also significantly increased the muscle fiber diameter and the capillary-to-muscle fiber ratio in gastrocnemius muscles compared with all other ischemic groups (P<0.05). CONCLUSION: These findings suggested that chitosan and MCs together could offer a new approach for the therapeutic induction of angiogenesis in cases of peripheral arterial diseases.

Histomorphometric and immunohistochemical evaluation of angiogenesis in ischemia by tissue engineering in rats: Role of mast cells.

The aim of this study was to find a proper method for improvement of ischemic condition in the rat hind limb and also to observe the efficacy of cell engraftment with alginate/gelatin three-dimensional scaffolds. Eighteen male Wistar rats weighing 200 to 250 g were randomly divided into three groups (n = 6) including a) ischemia group; in which femoral artery was removed after ligation at the distance of 5 mm, b) scaffold group; in which hydrogel scaffold was added to the site of transected femoral artery and c) test group; in which in addition to hydrogel scaffold, mast cells (MCs) were also added (1 × 106 cells). Analysis of capillary density, artery diameter, histomorphometric parameters and immunohistochemistry in transected location were done on day 14 after femoral artery transection. The average number of blood capillary was significantly higher in the test group than other groups. Also, the average number of medium and large blood vessels was significantly higher in the test group compared to ischemia and scaffold groups. Application of MCs through the use of hydrogel scaffolds (alginate/gelatin) can be considered as a new approach in the application of stem cells for therapeutic angiogenesis under ischemic conditions which can improve the angiogenesis process in patients with peripheral artery diseases.

Developing Rat Bone Marrow Derived Mast Cells by the Splenic Cells Culture Supernatant of Rat and Mouse.

BACKGROUND: Mast cells play a critical role in the pathogenesis of various immunological and non-immunological diseases. It is now accepted that culturing primary mast cells considered as a tool for investigation role of mast cells in diseases. Development of various animal primary mast cells and their function could be used for the translational studies in the pathogenesis of human diseases. The aim of the study was to develop simple and cost-efficient method for differentiation and culture of rat mast cells from bone marrow by using rat and mouse spleen supernatant. MATERIALS AND METHODS: Bone marrow cells from 10 to15-weeks-old male rats was obtained and cultured for three weeks on cell culture medium. After that, purity of cells was approved by FCɛRI and CD117 antibodies, toluidine blue and Immunohistochemistry (IHC). RESULTS: After 3 weeks continuous culturing, high purity of cells was found. CD117, CD34 expression and tryptase were 80.1, 76.89 and 87.9%, respectively by rat splenic supernatant, whereas 85.4, 83.07 and 82.1%, respectively with mouse splenic supernatants. Besides, rat spleen supernatant developed 91.4% and mouse splenocyte supernatant developed 89.7% mast cells based on surface markers. CONCLUSION: The data presented in this study indicated equal maturation and differentiation of bone marrow derived rat mast cells by using both spleen supernatants.

Role of Mast Cells and Type 2 Innate Lymphoid (ILC2) Cells in Lung Transplantation.

The multifunctional role of mast cells (MCs) in the immune system is complex and has not fully been explored. MCs reside in tissues and mucous membranes such as the lung, digestive tract, and skin which are strategically located at interfaces with the external environment. These cells, therefore, will encounter external stimuli and pathogens. MCs modulate both the innate and the adaptive immune response in inflammatory disorders including transplantation. MCs can have pro- and anti-inflammatory functions, thereby regulating the outcome of lung transplantation through secretion of mediators that allow interaction with other cell types, particularly innate lymphoid cells (ILC2). ILC2 cells are a unique population of hematopoietic cells that coordinate the innate immune response against a variety of threats including infection, tissue damage, and homeostatic disruption. In addition, MCs can modulate alloreactive T cell responses or assist in T regulatory (Treg) cell activity. This paper outlines the current understanding of the role of MCs in lung transplantation, with a specific focus on their interaction with ILC2 cells within the engrafted organ.