Transplantation of bone-marrow-derived mesenchymal and epidermal stem cells contribute to wound healing with different regenerative features

A new concept for wound therapy is the initiation of the regeneration of epidermal and dermal layers with appendages for skin function recovery. Bone-marrow-derived mesenchymal and epidermal stem cells (BMSCs and SSCs) are hypothesized to be able to home toward or to be transplanted to wound sites for skin repair and regeneration, but this awaits confirmation by further experimental and clinical evidence. In this study, the influence of the transplantation of BMSCs and SSCs with porous gelatin-β-tricalcium phosphate sponge as scaffolds on wound re-epithelization, collagen synthesis, skin tensile strength recovery, and skin appendage regeneration has been investigated. The transplantation of BMSCs or SSCs significantly accelerates wound re-epithelization, stimulates dermal collagen synthesis, and exhibits the trend to enhance the tensile strength recovery of skin. Furthermore, regenerative features of BMSCs and SSCs have been identified in activating blood vessel and hair follicle formation, respectively. These results not only provide experimental evidence for the application of BMSCs and SSCs as promising therapeutics for clinical wound treatment, but also display their characteristics in activating distinct skin appendage regeneration, which might have novel applications in skin tissue engineering.     

Defective macrophage function in wound repair of P/J mice

The strength of healing full-thickness incised dermal wounds in P/J mice was less than that of CD-1 mice although the strength of intact skin was similar for each strain. Five days after surgery, P/J mice had wounds with tensile strengths of 65 +/- 18g while CD-1 mice had wounds with strengths of 85 +/- 15g. The wound breaking strength of P/J mice was restored to normal values (86 +/- 18g) by administering glucan. The consequence of defective monocytes in wound repair is discussed in reference to P/J mice.     

miR-155 promotes cutaneous wound healing through enhanced keratinocytes migration by MMP-2

Inflammation, re-epithelization and tissue remodeling are three essential steps during wound healing. The re-epithelization process plays the most important role which mainly involves keratinocyte proliferation and migration. miR-155 has been reported to participate in cell migration and transformation, however, its function in skin wound healing is largely unknown. Here we hypothesize that overexpression of miR-155 at wound edges could accelerate wound healing mediated by enhanced keratinocyte migration. To test this hypothesis, direct local injection of miR-155 expression plasmid to wound edges was conducted to overexpress miR-155 in vivo. Results shown that miR-155 significantly promoted wound healing and re-epithelization compared to control, while did not affect wound contraction. Also, miR-155 overexpression accelerated primarily cultured keratinocyte migration in vitro, but had no effect on cell proliferation. Importantly, western blot analysis shown that MMP-2 was significantly upregulated whiles its inhibitor TIMP-1 downregulated after miR-155 treatment. Moreover, the use of ARP-101, an MMP-2 inhibitor, effectively attenuated the accelerative effects on cell migration induced by miR-155. Taken together, our results suggest that miR-155 has the promote effect on wound healing that is probably mediated by accelerating keratinocyte migration via upregulated MMP-2 level. This study provides a rationale for the therapeutic effect of miR-155 on wound healing.     

Impaired wound healing in transgenic mice overexpressing the activin antagonist follistatin in the epidermis

Recently, we demonstrated a strong upregulation of activin expression after skin injury. Furthermore, overexpression of this transforming growth factor beta family member in the skin of transgenic mice caused dermal fibrosis, epidermal hyperthickening and enhanced wound repair. However, the role of endogenous activin in wound healing has not been determined. To address this question we overexpressed the soluble activin antagonist follistatin in the epidermis of transgenic mice. These animals were born with open eyes, and the adult mice had larger ears, longer tails and reduced body weight compared with non-transgenic littermates. Their skin was characterized by a mild dermal and epidermal atrophy. After injury, a severe delay in wound healing was observed. In particular, granulation tissue formation was significantly reduced, leading to a major reduction in wound breaking strength. The wounds, however, finally healed, and the resulting scar area was smaller than in control animals. These results implicate an important function of endogenous activin in the control of wound repair and scar formation.     

Effect of exogenous fibronectin on the healing of skin wounds as evidenced by tensiometric data

Fibronectin (FN) solution purified from human blood plasma significantly increased the breaking strength of skin wounds in rats, when applied locally and/or intravenously. The best effect (more than 2-fold) was revealed after combined local and i. v. application of FN. Maximum stimulating activity was observed on the 3rd postoperative day, directly indicating the importance of FN for the early phases of wound healing. The data suggest the possibility of using FN preparations for the stimulation of tissue repair processes.     

Nerve dependency in scarless fetal wound healing

The human fetus is capable of healing cutaneous wounds without scar up to the third trimester of development This process of tissue repair is more akin to newt limb regeneration than classic adult scar forming wound repair. Regeneration of the newt limb is dependent on neural input in its early stages. This study was an attempt to determine whether a similar dependence on neural input exists for mammalian fetal wounds to heal without scar. The left hind limb of six fetal lambs was denervated during the early second trimester of development (day 55; term = 145 days). Two weeks after denervation, the animals were again exposed to create bilateral incisional and 6-mm-diameter excisional wounds on their innervated right and denervated left lower extremities. Five days after creation of these defects, the wounds were examined for alterations in repair. Four fetal lambs survived, and three were suitable for evaluation. There were marked alterations in wound healing seen after denervation. Excisional wounds on the innervated side contracted and decreased their surface area by 14 percent. In contrast, the denervated wounds not only failed to contract, but increased in size by 60 percent. Changes in the incisional wounds were equally distinctive. Innervated incisional wounds healed completely without scar and had a wound breaking strength comparable to that of normal skin (Table I). In contrast, two of the three denervated incisional wounds dehisced and failed to heal, even in the regions where the skin was approximated by suture. The third denervated incisional wound did heal but with a significant amount of scar. Electron microscopy confirmed this finding by clearly demonstrating thickened and irregular collagen deposition in the extracellular matrix of all the denervated incisional specimens. In summary, like the regenerating newt limb, scarless fetal skin wound repair requires neural stimulation for tissue regeneration to occur. Therefore, in the mammal, the primary regulator for this unique type of tissue repair may have a central neural, rather than a local, tissue origin.     

Vanadate ingestion increases the gain in wound breaking strength and leads to better organized collagen fibers in rats during healing

Repair of incision wounds closed by suturing is evaluated by the progressive gain in wound breaking strength. Previously the closure of open wounds in rats ingesting vanadate, an inhibitor of tyrosine phosphate phosphatases, was shown to occur with deposition of more uniformly organized collagen fiber bundles. The hypothesis of this study was that deposition of more uniformly organized collagen fibers would enhance the gain in wound breaking strength of incisional wounds. Six adult rats received vanadate-supplemented saline drinking water for 1 week before placement of two 6-cm, parallel, suture-closed wounds on their backs. Six control rats received identical wounds and were given saline drinking water. The drinking water regimen was continued for 1 week after wounding, and then wound strength was tested with a tensiometer and tissue samples were obtained for histologic evaluation. Wound breaking strength doubled in vanadate-treated rats compared with controls. Bright-field and polarized light microscopy showed that the connective tissue matrix of granulation tissue from control rats was oriented perpendicular to the surface of the skin. In contrast, the connective tissue matrix of granulation tissue from vanadate-treated rats was oriented parallel to the skin surface. The gap in granulation tissue between the edges of the wounds in the vanadate-treated rats was greater than that in controls. Electron microscopy showed that wounds in the vanadate-treated contained uniform collagen fibers that were 20 percent greater in diameter and more evenly spaced than they were in controls. It is proposed that these changes in the organization of collagen fibers within incisional wounds were responsible for the increased wound breaking strength observed in rats ingesting vanadate.     

Hyperbaric Oxygen Therapy to Treat Diabetes Impaired Wound Healing in Rats

Wound healing in diabetes is frequently impaired and its treatment remains a challenge. Hyperbaric oxygen therapy (HBOT) receives a wide attendance and is often used as a last resort treatment option, however, its effectiveness for many conditions is unproven. We tested the effect of HBOT on healing of diabetic ulcers in an animal experimental setting. Experimental diabetes was induced by intraperitoneal injection of streptozotocin. Four weeks after diabetes induction, rats were ulcerated by clamping a pair of magnet disks on the dorsal skin for 16 h. After magnet removal, the animals received HBOT, daily on weekdays, for 4 weeks. To examine the effect of HBOT on diabetes impaired wound healing, the degree of wound tissue perfusion, inflammation, angiogenesis, and tissue breaking strength were evaluated. HBOT effects on the degree of inflammation and number of blood vessels could not be observed. HBOT improved the tissue breaking strength of the wound, however, this did not reach statistical significance. Twenty hours after ending the HBOT, a significantly improved oxygen saturation of the hemoglobin at the venous end of the capillaries and the quantity of hemoglobin in the micro-blood vessels was measured.     

Effects of procoagulants on wound healing

As blood coagulation is a prelude for wound healing, a systemic haemocoagulant (Botropase) and local procoagulants (thrombin and fibrin) were evaluated on physical (wound breaking strength, wound half-closure time and period of epithelization), biochemical (granuloma-hydroxyproline and hexosamine) and histological attributes of healing wounds in albino rats. Botropase prompted all phases of tissue repair. Thrombin delayed wound contraction whereas fibrin had no discernable action. The findings that procoagulants modify healing process has bearing on their surgical use.     

Hydroalcoholic extract based-ointment from Punica granatum L. peels with enhanced in vivo healing potential on dermal wounds

The present study reports for the first time, the in vivo wound healing potential of Punica granatum L. peels. A 5% (w/w) methanolic extract based-ointment was formulated and evaluated for its wound healing in guinea pigs. The ointment was applied in vivo on the paravertebral area of twelve excised wounded models once a day for 10 consecutive days. The ointment significantly enhanced the wound contraction and the period of epithelialization as assessed by the mechanical (contraction rate, tensile strength), the biochemical (increasing of collagen, DNA and proteins synthesis) and the histopathological characteristics. Such investigation was encouraged by the efficiency of the methanolic extract as antimicrobial and antioxidant. Indeed, the extract showed antioxidant activity as strong as natural and synthetic compounds (Trolox, BHA, Quercetin). Furthermore, the extract exhibited significant antibacterial and antifungal activity against almost all tested bacteria: Pseudomonas aeruginosa ATCC 9027, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, Klebsiella pneumoniae, Salmonella anatum, Salmonella typhimurium, Streptococcus pneumoniae, and fungi Candida albicans, Candida glabrata, Trichopyton rubrum and Aspergillus niger. The formulated ointment might well find use as skin repair agent without hazard to human health based on these results and on the fact that it has been well established that the extracts of pomegranate used in conditions similar to those applied by traditional medicine, showed no toxic effects.     

Activation of the EPOR-β common receptor complex by cibinetide ameliorates impaired wound healing in mice with genetic diabetes

Diabetes is characterized by poor wound healing which currently lacks an efficacious treatment. The innate repair receptor (IRR) is a master regulator of tissue protection and repair which is expressed as a response injury or metabolic stress, including in diabetes. Activation of the IRR might provide benefit for diabetic wound healing. A specific IRR agonist cibinetide was administered in an incisional wound healing model performed mice with genetic diabetes (db⁺/db⁺) and compared to the normal wild-type. Animals were treated daily with cibinetide (30 μg/kg/s.c.) or vehicle and euthanized 3, 7, and 14 days after the injury to quantitate vascular endothelial growth factor (VEGF), malondialdehyde (MAL), phospho-Akt (pAkt), phospho e-NOS (p-eNOS), and nitrite/nitrate content within the wound. Additional evaluations included quantification of skin histological change, angiogenesis, scar strength, and time to complete wound closure. Throughout the wound healing process diabetic animals treated with vehicle exhibited increased wound MAL with reduced VEGF, pAkt, peNOS and nitrite/nitrate, all associated with poor re-epitheliziation, angiogenesis, and wound breaking strength. Cibenitide administration significantly improved these abnormalities. The results suggest that cibinetide-mediated IRR activation may represent an interesting strategy to treat diabetes-associated wound healing.     

Essential fatty acids are not required for wound healing.

Rats with essential fatty acid deficiency (EFAD) exhibit mild body growth retardation, diminished leukocyte influx in certain models of inflammation, and skin lesions characterized by ulceration, thinning and decreased pigmentation. In the present study we examined the role of EFAD in cutaneous wound healing, a process in which the inflammatory response and the macrophage play a central role. We reproduced the EFAD condition in Lewis rats (n = 35), and examined its effects in wound healing using the paired rat surgical incision model. Rats were compared with weight-matched controls, receiving standard chow diet. Skin samples harvested at days 5, 7, 14 and 21 post-wounding were evaluated for tensiometry and histology. EFAD rats exhibited all the characteristics of this condition, and the typical alteration of liver lipids. Skin samples harvested at different days post-wounding did not show difference in maximal breaking strength when compared to weight-matched controls. Histological evaluation of skin samples showed no difference in the cellular inflammatory infiltration in either EFAD rats or in weight-matched controls. Immunohistochemical studies revealed no difference in the influx of macrophages in the different groups of rats. Fatty acid supplementation of EFAD rats (n = 7), successfully reversed the EFAD state as assessed by the macroscopic skin and liver changes and liver fatty acid content, without modifying either tensile strength or cellular inflammatory infiltration. Our results suggest that EFAD does not alter the normal course of the cutaneous wound repair in rats, despite all the cutaneous alterations produced by this condition. We conclude that essential fatty acids (EFAs) are not essential for cutaneous wound repair.     

角蛋白/海藻酸钠/聚丙烯酰胺水凝胶皮肤敷料的制备及创口修复研究

为了获得更为理想的皮肤创口修复敷料,在海藻酸钠(SA)和聚丙烯酰胺(PAM)水凝胶的基础上复合人发角蛋白(KTN),制得KTN/SA/PAM水凝胶皮肤敷料.用电子万能测试机、扫描电子显微镜等对其进行表征,结果显示,KTN/SA/PAM水凝胶皮肤敷料拉伸强度为42.41 kPa,弹性模量11.19 kPa,接近人体皮肤组...     

Studies on wound healing potential of red pigment isolated from marine Bacterium Vibrio sp.

Wounds are common clinical entities of life which may be subacute or acute. Wound healing is a complex biochemical process where the cell structures are restored to normalcy, which depend on cell proliferation and migration, basically fibroblast cell. The present investigation was undertaken to evaluate the healing efficacy of red pigment isolated from marine isolate Vibrio sps on experimental wounds in albino rats. The red pigment was applied topically, twice daily for 14 days. Treatment with framycetin ointment was used as reference control. The red pigment treated group showed faster reduction in wound area in comparison with control and framycetin ointment treated groups. In conclusion, red pigment possesses significant healing potential in wounds and has a positive influence on the different phases of wound repair.     

Wound healing property of ethanolic extract of leaves of Hyptis suaveolens with supportive role of antioxidant enzymes

Ethanolic extract of leaves of Hyptis suaveolens was evaluated for its wound healing activity in ether-anaesthetized Wistar rats at two different doses (400 and 800 mg/kg) using incision, excision, and dead space wound model. Significant increase in skin breaking strength, granuloma breaking strength, wound contraction, hydroxyproline content and dry granuloma weight and decrease in epithelization period was observed. A supportive study made on granuloma tissue to estimate the levels of catalase and superoxide dismutase recorded a significant increase in the level of these antioxidant enzymes. Granuloma tissue was subjected to histopathological examination to determine the pattern of lay-down for collagen using Van Gieson and Masson Trichrome stains. Enhanced wound healing activity may be due to free radical scavenging action of the plant and enhanced level of antioxidant enzymes in granuloma tissue. Better collagenation may be because of improved antioxidant studies.     

Copper-induced change in the ESR signal of hemoglobin nitrosyl complexes in wound by the action of copper nanoparticles

The results concerning changes in the ESR signal of hemoglobin nitrosyl complexes in wound tissues in the course of healing by the action of ointments with copper nanoparticles (patent N2460532, Russia) are presented. It is shown that the wound healing process modified by the influence of copper nanoparticles demonstrates the increase in the ESR signal amplitude for hemoglobin nitrosyl complexes as compared with controls (the ointment base without nanoparticles). Planimetric measurements of wound area through reparation course indicate an active process of wound healing for injuries treated with copper nanoparticles in the ointment, resulting in lessening half-reparation time up to 5.0 times as compared with controls (treatment with the ointment base). The paper discusses the role of copper nanoparticles, NO and their potential synergistic effect on the skin wound regeneration.     

Novel Clotrimazole and Vitis vinifera loaded chitosan nanoparticles: Antifungal and wound healing efficiencies

Chitosan integrated nanoparticles of clotrimazole and Egyptian Vitis vinifera juice extract was evaluated in order to maximize the antifungal activity and reduce the gross side effects. In the present study Egyptian Thompson Seedless Vitis vinifera and Clotrimazole (Cz) loaded chitosan nanoparticles (NCs/VJ/Cz) showed a promising antifungal effect with average inhibition zone diameters of 74 and 72 mm against Candida albicans and Aspergillus niger respectively. NCs/VJ /Cz was stable with significant drug entrapment efficiency reached 94.7%; PDI 0.24; zeta potential value + 31 and average size 35.4 nm diameter. Ex vivo and in vivo evaluation of skin retention, permeation and wound repair potentialities of NCs/VJ /Cz ointment was examined by experimental rats with wounded skin fungal infection. Data proved the ability of NCs/VJ /Cz to gradually release the drugs in a sustained manner with complete wound healing effect and tissue repair after 7 days administration. As a conclusion NCs/VJ /Cz ointment can be used as a novel anti-dermatophytic agent with high wound healing capacity.     

The antagonism of glucocorticoid inhibition of wound healing in rats by growth hormone-releasing factor

Daily therapeutic injections of cortisone to rats will cause weight loss and impaired wound healing. Weight loss is attributed to the catabolic effect of steroid, whereas impaired healing is associated with reductions in fibroplasia and connective tissue deposition. As the major structural protein component of connective tissue is collagen, its absence is responsible for the retarded gain in wound breaking strength. Cortisone also blocks wound closure by inhibiting wound contraction. An anabolic agent such as growth hormone may antagonize the effect of cortisone on the wound healing process. Endogenous GH can be released from the pituitary by exogenous injections of growth hormone-releasing factor (GRF). Two synthetic GRF peptides, a natural 44-amino acid peptide of the human GRF sequence, GRF-44, and an N-terminally substituted analog 29 residues, GRF-29A, were studied. Each was given twice daily with a single daily injection of cortisone for a 7-day period. Concurrent administration of GRF-44 or GRF-29A and cortisone to rats had no effect on restored body weight loss or inhibited wound contraction. While GRF-44 restored collagen deposition and caused restored wound breaking strength, GRF-29A was ineffective in restoring either. GRF-44, a synthetic peptide that stimulates pituitary release of growth hormone, antagonized some of the inhibiting effect of steroid on wound repair by promoting fibroplasia and collagen deposition.     

Assay of radiation effects in mouse skin as expressed in wound healing

The effect of 150 kVp X irradiation on the healing of full depth surgical wounds in the lower dorsal skin of the mouse was assayed by measuring the wound strength of seven 2-mm-wide segments along each wound. The strength of unirradiated wounds increased with time in two phases: during the first 2 weeks it reached nearly half of the values recorded from unwounded skin, after which the rate of increase slowed for at least 2 weeks before beginning a second increase. By 150 days, the breaking strength of the wound was about 80% of that of unwounded skin. A single dose of 18 Gy prior to wounding reduced the strength of the wounds to about one-third to one-half that of an unirradiated wounds within the 3 months of follow-up. The effect of irradiation on wound strength did not change as the interval between exposure and wounding was increased to 2 months but decreased slightly when this interval was extended to 3 months. When the healing wound was irradiated within 5 days of surgery, the effect on healing was about the same as with preirradiation; if irradiation was delayed for 12 days after wounding the second phase of healing was only postponed and the wound strength ultimately approached the values recorded from unirradiated wounds. The wound strength of skin preirradiated by X rays and assayed 14 days after wounding showed a clear sigmoid dose response with a threshold between 8 and 10 Gy and a plateau at the maximum effect above 20 Gy. The persistence for at least 3 months of the effect of radiation on wound healing suggests that the tissues involved in the healing process are normally proliferating slowly. The accelerated expression of radiation injury through surgical wounding permits the early quantification of the radiation response of tissues that would normally be delayed in their expression of radiation damage.     

上海交通大学医学院附属新华医院崇明分院检验科

目的:探讨富血小板血浆(platelet-rich plasma,PRP)结合组织工程皮肤对裸鼠巨大创面修复的影响。方法:首先,利用密度梯度离心法制备富含生长因子的浓缩血小板的血浆,并测其所含生长因子的量;其次,在裸鼠的背侧部分构建大面积皮肤创面,分别用人工真皮,组织工程皮肤,碱性成纤维细胞生长因子组织工程皮肤,表皮生长因子组织工程皮肤和PRP结合组织工程皮肤修复裸鼠巨大创面;最后,手术后不同时间间隔收集组织标本,采用HE染色,PAS染色和免疫组化等方法评估创面愈合情况。结果:PRP结合组织工程皮肤组创面修复愈合情况最好。