Current regenerative medicine-based approaches for skin regeneration: A review of literature and a report on clinical applications in Japan.

Current trends indicate a growing interest among healthcare specialists and the public in the use of regenerative medicine-based approaches for skin regeneration. The approaches are categorised in either cell-based or cell-free therapies and are reportedly safe and effective. Cell-based therapies include mesenchymal stem cells (MSCs), tissue induced pluripotent stem cells (iPSCs), fibroblast-based products, and blood-derived therapies, such as those employing platelet-rich plasma (PRP) products. Cell-free therapies primarily involve the use of MSC-derived extracellular vesicles/exosomes. MSCs are isolated from various tissues, such as fat, bone marrow, umbilical cord, menstrual blood, and foetal skin, and expanded ex vivo before transplantation. In cell-free therapies, MSC exosomes, MSC-derived cultured media, and MSC-derived extracellular vesicles are collected from MSC-conditioned media or supernatant. In this review, a literature search of the Cochrane Library, MEDLINE (PubMed), EMBASE, and Scopus was conducted using several combinations of terms, such as 'stem', 'cell', 'aging', 'wrinkles', 'nasolabial folds', 'therapy', 'mesenchymal stem cells', and 'skin', to identify relevant articles providing a comprehensive update on the different regenerative medicine-based therapies and their application to skin regeneration. In addition, the regulatory perspectives on the clinical application of some of these therapies in Japan are highlighted.

Treatment of bronchial fistula after extraplural pneumonectomy using flexible bronchoscopy with the administration of OK432, fibroblast growth factor basic and fibrin glue sealant.

Treatment options for bronchial fistula (BF) after pneumonectomy are often limited and carry significant morbidity and mortality. The patient underwent right extrapleural pneumonectomy for malignant pleural mesothelioma had BF without macroscopic fistula found by bronchography. We treated this minor BF using bronchoscopy with the administration of OK-432, fibroblast growth factor basic, and fibrin glue sealant. Two weeks after this treatment, we confirmed the improvement of the fistula by bronchography. Bronchoscopic therapy for BF was useful for a small, early fistula without infection.

Laparoscopic surgery for colorectal cancer is not associated with an increase in the circulating levels of several inflammation-related factors.

It has been hypothesized that inflammatory response triggered by surgery might induce the release of molecules that could promote proliferation, invasion and metastasis of surviving cancer cells. To test this hypothesis, the levels of multiple inflammation-related circulating factors were analyzed in patients undergoing surgery for colorectal cancer. A Luminex xMAP system was used to simultaneously assess levels of IL-1ß, IL-1ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, IL-17, FGF, eotaxin, G-CSF, GM-CSF, IFN-γ, IP-10, MCP-1, MIP-1α, MIP-1ß, PDGF-BB, RANTES, TNF-α and VEGF in 20 colorectal cancer patients and 10 age-matched non-neoplastic patients. In cancer patients analyses were performed at baseline (before surgery) and at different time points (up to 30 days) following laparoscopic surgery. Significantly higher levels of IL-1ß, IL-7, IL-8, G-CSF, IFN-γ and TNF-α were detected in colorectal cancer patients compared to controls at baseline. In colorectal cancer patients, circulating levels decreased progressively following surgery and after day 30 post-surgery were no longer different from controls. These findings suggest that expression levels of several cytokines are higher in colorectal cancer patients compared to control subjects and no significant increase in several inflammation-related circulating factors is observed following laparoscopic surgery for cancer. Confirmation and validation in a different and larger cohort of patients are warranted.

Effect of fibroblast growth factor and enamel matrix derivative treatment on root resorption after delayed replantation.

BACKGROUND: Periodontal ligament (PDL) healing and long term prognosis of replanted avulsed teeth should rely on several factors including length of extra-oral dry time and type of the storage medium. The status of periodontal ligament is critical for the healing of replanted teeth. Different substances have been used for root surface treatment to promote formation of PDL and increase the survival of avulsed teeth submitted to replantation. AIM: The purpose of this study was to assess the effect of recombinant basic fibroblast growth factor 2 (bFGF) and enamel matrix derivative (EMD) on root resorption after delayed replantation. DESIGN: 18 freshly extracted single-rooted incisor and premolar teeth were extracted from the beagle dogs and immersed in whole bovine milk for 45 and 60 min (n = 3 each). Following storage period, sockets washed and teeth were treated with bFGF and EMD and replanted into the sockets. After 8 weeks, dogs were sacrificed, specimens processed to 4-µm thick serial sections for histopathologic examination and morphometric assessments. Thus, the proportions of the roots that exhibited signs of surface resorption, inflammatory resorption, and replacement resorption, that is, ankylosis and normal PDL were noted. RESULTS: The percentage of root resorption was in the following order: EMD>milk>bFGF for 45 min and milk>EMD>bFGF for 60 min. For all groups, teeth stored 60 min showed significantly higher incidence of PDL resorption than those stored for 45 min (P < 0.01). The highest incidence of replacement resorption was observed in teeth treated with EMD for 60 min. After 8 weeks, the least resorption was found in bFGF-treated group (P < 0.01). CONCLUSIONS: The findings of this study suggest that use of bFGF favored the formation of new periodontal ligament; prevent ankylosis and resorption process following delayed replantation of teeth while EMD shows replacement resorption, which may turn to ankylosis.

Age associated communication between cells and matrix: a potential impact on stem cell-based tissue regeneration strategies.

A recent paper demonstrated that decellularized extracellular matrix (DECM) deposited by synovium-derived stem cells (SDSCs), especially from fetal donors, could rejuvenate human adult SDSCs in both proliferation and chondrogenic potential, in which expanded cells and corresponding culture substrate (such as DECM) were found to share a mutual reaction in both elasticity and protein profiles (see ref. (1) ). It seems that young DECM may assist in the development of culture strategies that optimize proliferation and maintain "stemness" of mesenchymal stem cells (MSCs), helping to overcome one of the primary difficulties in MSC-based regenerative therapies. In this paper, the effects of age on the proliferative capacity and differentiation potential of MSCs are reviewed, along with the ability of DECM from young cells to rejuvenate old cells. In an effort to highlight some of the potential molecular mechanisms responsible for this phenomenon, we discuss age-related changes to extracellular matrix (ECM)'s physical properties and chemical composition.

Ability of plasmid DNA complexed with histidinylated lPEI and lPEI to cross in vitro lung and muscle vascular endothelial barriers.

DNA complexes made with cationic polymers (polyplexes) developed as nonviral vectors for gene therapy must be enabled to cross through vascular endothelium to transfect underlying tissues upon their administration in the blood circulation. Here, we evaluated the transendothelial passage (TEP) of DNA complexes made with histidinylated linear polyethylenimine (His-lPEI) or linear polyethylenimine (lPEI). In vitro studies were performed by using established transwell lung and skeletal muscle vascular endothelial barriers. The models were composed of a monolayer of human lung microvascular endothelial (HMVEC-L) cells and mouse cardiac endothelial (MCEC) cells formed on a PET insert and immortalized human tracheal epithelial (ΣCFTE29o-) cells and mouse myoblasts (C2C12) as target cells cultured in the lower chamber, respectively. When the vascular endothelium monolayer was established and characterized, the transfection efficiency of target (ΣCFTE29o- and C2C12) cells with plasmid DNA encoding luciferase was used to evaluate TEP of polyplexes. The luciferase activities with His-lPEI and lPEI polyplexes compared to those obtained in the absence of endothelial cell monolayer were 6.5% and 4.3% into ΣCFTE29o- cells, and 18.5% and 0.23% into C2C12 cells, respectively. The estimated rate for His-lPEI polyplexes was 0.135 µg/cm(2).h and 0.385 µg/cm(2).h through the HMVEC-L and MCEC monolayers, respectively. These results indicate that His-lPEI polyplexes can pass through the lung and skeletal muscle vascular endothelium and can transfect underlying cells.

Acupuncture on the Basic Fibroblast Growth Factor and Type Ⅰ Collagen in Colons of Rats with Crohn's Disease / 针灸推拿医学（英文版）

Objective:To observe the impacts of herb-partitioned moxibustion,warm moxibustion and electroacupuncture on the basic fibroblast growth factor(bFGF)and collagen type Ⅰ(Col Ⅰ)in colons of rats with Crohn' disease(CD),and discuss the mechanism of acupuncture therapy on the intestinal fibrosis in CD.Methods:The model rats were developed by TNBS as multiple proinflammatory method.The rats were randomly divided into 5 groups:a normal group,a model group,a warm moxibustion group,an electroacupuncture group and a herb-partitioned moxibustion group.The treatments were carried out at Tianshu(ST 25)(bilateral)and Qihai(CV 6)in different treatments.The immunohistochemistry was used to detect the expression position of Col Ⅰ and bFGF.Results:The expressions of Col Ⅰ and bFGF in colons of rots in the model group significantly increased(compared with the normal group,P<0.01).After the herb-partitioned moxibustion,warm moxibustion and electroacupuncture,the expressions of Col Ⅰ and bFGF reduced markedly in the rats with CD(P<0.01).The expression of bFGF and Col Ⅰ in the colons had an obvious correlation in the Spearman rank correlation analysis.Conclusion:Acupuncture treatment reduced the abnormally high levels of expressions for Col Ⅰ and bFGF in colons.Col Ⅰ and bFGF participated in the fibrosis.Acupuncture treatment may reduce the bFGF expression in colons to regulate the excessive deposition,treating the intestinal fibrosis in CD.

The experimental study of bFGF-fibrin gule-amniotic membrane on corneal alkali burns / 重庆医科大学学报

Objective:To study the effect of a new medical biomaterial—bfgf-fibrin glue-amniotic membrane(bFAM) for corneal alkali burn and provide experimental foundation for the clinical application in the future. Methods: We use fibrin glue compounded with bFGF and amniotic membrane to prepare the bFAM. 45 New Zealand rabbits were used to make corneal alkali burn models and divided into 3 groups:bFAM group、amniotic membrane group、comparison group, each group have 15 rabbits. After operation, we observe the linical curative effect. 7d、14d and 28d after operation, 5 corneas were extracted for histopathological analysis. The expression of bFGF in corneas was measured by immunohistochemistry analysis. Results: After operation,bFAM group have better curative effect than amniotic membrane group in Inflammatory reaction、tissue repair and anti-CNV. The expression of bFGF in bFAM group is much higher than amniotic membrane group and comparison group in the day 7 afer operation, and in other time points the expression have not distinct difference in all groups. Conclusion: Compared with single layer amniotic membrane, bFAM shows better curative effect for corneal alkali burn.

Expression of Fas/FasL ligand and cell apoptosis in ischemia/reperfusion-induced retina and effects of bFGF / 国际眼科杂志(Guoji Yanke Zazhi)

· AIM: To explore the relationship between the expression of Fas/FasL and the apoptosis in retinal ischemia/reperfusion injury of rats, as well as the therapeutic effects of basic fibroblast growth factor (bFGF)on the ischemic retina.injury were made by transiently elevating introcular pressure. A total of 28 rats were divided into Normal Group and Operative Group. The latter were subdivided into 1, 6, 12, 24, 48 and 72h after reperfusion, in which the left eyes of the rats were in the ischemia/reperfusion groups and the right ones were in the treatment groups (bFGF intracameral injection). Apoptosis was assessed by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labelling (TUNEL) method, and theexpression of Fas/FasL ligand was studied by strept avidin-biotin complex (SABC) immunohistochemistry.mai rats' retinae, but there were a significant number of TUNEL positive cells in 6-24h after transient ischemia followed by a decrease at 48h. The number of TUNEL positive cells reached a maximum at 24h after ischemia.The expression of Fas gradually increased as early as at 6h, reached a peak at 24h, then decreased at 48h. Similarly, the expression of Fas ligand was at peak in 24-48h in GCL and INL of retina. bFGF ministered before reperfusion inhibited apoptotsis and ameliorated the tissue damage. It also diminished Fas and FasL expression in ischemic/reperfused retina.siently elevated IOP induced apoptosis of cells in the retina. Fas/FasL may have an important role in the early events of the apoptotic pathways. bFGF can rescue RGCs from retinal ischemia/reperfusion injury through down-regulation of Fas and Fas ligand expression and may represent an important mechanism for therapeutic neuroprotection.

Effects of scallop skirt glycosaminoglycan on proliferation of vascular smooth muscle cells in rats / 中国药理学与毒理学杂志

AIM To investigate if scallop (Placopecta magellanicus) skirt glycosaminoglycan (SS-GAG) inhibits the proliferation of vascular smooth muscle cell (VSMC) as heparin does so and to clarify its mechanism. METHODS The inhibitory effects of SS-GAG on the proliferation of rat thoracic aorta and abdominal aorta VSMC induced by fetal bovine serum (FBS) or basic fibroblast growth factor (bFGF) were determined by cell counting, crystal violet staining and MTT colorimetry. The effects of SS-GAG on the expression of proliferating cell nuclear antigen (PCNA) and platelet-derived growth factor (PDGF) in VSMC proliferation induced by bFGF were evaluated by immunohistochemical technique (LSAB method) and computer image analysis system. RESULTSSS-GAG exerted antagonistic effects on VSMC proliferation induced by 20% FBS and 50 μg·L-1 bFGF at concentrations ranging from 50 mg·L-1 to 200 mg·L-1 and repressed the increasing expression of PCNA and PDGF. CONCLUSION SS-GAG significantly inhibits the proliferation of VSMC, which may be carried out through repression of PDGF and PCNA expression.

EXPLORATION OF THE MECHANISM OF THERAPEUTIC EFFECT OF BASIC FIBROBLAST GROWTH FACTOR IN THE TREATMENT OF BLAST-INDUCED DEAFNESS / 解放军医学杂志

To explore the mechanism of therapeutic effect of basic fibroblast growth factor (bFGF) in the treatment of blast-induced deafness, and to define its optimal clinical use, bFGF was infused into the guinea pig's cochlea, combined with intramuscular injection of bFGF after being exposed to explosion. The compound action potential (CAP) and auditory brainstem response (ABR) were measured in these animals. 125I labeled basic fibroblast growth factor ( 125I -bFGF) was injected intraperitoneal to the guinea pigs to observe whether it could pass through the blood-labyrinthine barrier. The results showed that bFGF infused to the cochlea might facilitate recovery of hearing loss following acoustic trauma. Basic fibroblast growth factor ( 125I -bFGF) intraperitoneally injected, could not pass through the blood-labyrinthine barrier. However intramuscular bFGF promoted the recovery of hearing, probably indirectly through the neuro-immunity network.

THE EFFECT OF LOCAL APPLICATION OF BFGF AND SUCRALFATE COMBINED WITH CTE ON VIABILITY AND STRETCH-BACK OF EXPANDED SKIN FLAP IN A PIG MODEL / 解放军医学杂志

To investigate the effect of bFGF and sucralfate on the improvement of the quality of expanded skin flap, white piglets were employed to establish a continuous tissue expansion model. They were randomly divided into 3 groups: group 1: both bFGF and sucralfate were injected; group 2: both bFGF and normal saline was injected; and group 3(the control group):only normal saline was injected. Three days after completion of the expansion ,normal and expanded skin flaps were created at random to assess flap viability and stretch back .The results showed that the flap survival length in group 1 was significantly larger than that of the control group and normal random flap( P

EFFECTS OF TRANSPOSITION OF FLAPS TO THE EXPRESSION OF bFGF IN CONCUSSION AREAS OF GUNSHOT INJURY / 解放军医学杂志

Flap pedicled with the arteria epigastrica superficialis was transposed to primarily repair firearm wound after the rabbit's posterior limb was shotted. Recovery of the wound was observed and the content of bFGF mRNA in myoideum of the concussion area was measured by reverse transcroption polymerase chain reaction(RT PCR). All wounds repaired with transposition of fascia flaps attained primary healing and the content of bFGF mRNA in myoideum of the concussion area was higher than that in the control group. The result suggests that transposition of arterialized fascia flaps could primarily repair firearm wounds successfully and the high expression of bFGF is one of the causes.

THE EXPERIMENTAL AND CLINICAL STUDY OF REPAIR OF BONE DEFECT BY MEANS OF TRANSPLANTING OSTEOBLAST AND BASIC FIBROBLAST GROWTH FACTOR / 解放军医学杂志

To study the method and the optimal quantity of osteoblast transplantation to stimulate bone regeneration by transplanting 6～8 day old cultured osteoblasts and bFGF, together with osteoblast free calvaria of fetal rabbits into the defects of the rabbit radius. The fibroblasts were embedded in the type Ⅰcollagen in the concentrations of 10 7 ,10 6 ,10 5 , 10 4 /ml. Bone repair was evaluated by X ray, bone density,and scanning electron microscopy. Bone union ratio of the radius with transplantation of 10 7 ,10 6 ,10 5 , 10 4 /ml osteoblasts was 40%, 65%, 31 25%, 5 55% respectively at 4 weeks after transplantation, and 41 67%, 75%, 37 5%, 10% respectively at 8 weeks after transplantation. Bone density of radius defect with transplantation of 10 7 , 10 6 , 10 5 , 10 4 /ml osteoblasts was (0 112?0 018), (0 159?0 033), (0 122?0 039), (0 066?0 002) respectively at 6 weeks after transplantation and (0 150?0 059), (0 173?0 041), (0 145?0 023), (0 103?0 023) respectively at 8 weeks after transplantation, In 10 cases of bone non union, union was achieved by transplanting fetal osteoblast and basic fibroblast growth factor. This finding indicated that that transplantation of fetal osteoblasts could stimulate bone defect repair. The optimal concentration of osteoblasts implant was 10 6 cells /ml. It might be used in the treatment of bone non union.

Expression of Fas and Fas ligand and cell apoptosis in the ischemia/reperfusion-induced retina and therapeutic effects of basic fibroblast growth factor / 中华眼底病杂志

Objctive To explore the relationship between the expression of Fas/FasL and the apoptosis occurs in retinal ischemia/reperfusion injury of rats, as well as the therapeutic effects of bFGF on the ischemic retina. Methods The models of retinal ischemia/reperfusion injury was made by transient elevating introcular pressure. A total of 28 rats were divided into normal and operation group.The latter were subdivided into 1 hour, 6, 12, 24, 48 and 72 hours after reperfusion group, in which the left eyes of the rats were in the ischemia/reperfusion groups and the right ones were in the treatment groups (bFGF intracameral injection). Apoptosis was assessed by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labelling (TUNEL) method, and the expression of Fas and Fas ligand was studied by strept avidin-biotin complex (SABC)immunohistochemistry. Results No positive cells were observed in the normal rats′ retinae, but there was a significant number of TUNEL positive cells in 6-24 hours after transient ischemia followed by a decrease at the 48th hour. The number of TUNEL positive cells reached a maximum at the 24th hour after ischemia. The expression of Fas gradually increased as early as when it was at the 6th hour, reached a peak at the 24th hour, and then decreased at the 48th hour. Similarly, the expression of Fas ligand was at peak in 24-48 hours in GCL and INL of retina. Conclusions Retinal ischemia-reperfusion after transient elevated IOP induced apoptosis of cells in the retina. Fas/FasL may play an important role in the early events of the apoptotic pathways. bFGF can rescue RGCs from retinal ischemia/reperfusion injury through downregulation of the expression of Fas/FasL and may represent an important mechanism for therapeutic neuroprotection.

Human mesenchymal stem cells differentiate into neuron-like cells with bFGF / 中华神经科杂志

Objective To investigate the differentiation from human mesenchymal stem cells (hMSC) into neuron-like cells. Methods hMSC were separated from rib marrow with Ficoll-Paque reagent and expanded in culture medium. hMSC were induced to differentiate into neurons with DMEM/BHA/DMSO or DMEM/monothioglycerol, respectively. Neuron-specific enolase (NSE), neurofilament (NF), nestin, glial fibrillary acidic protein (GFAP) were detected by immunohistochemistry. Results hMSC were expanded to be undifferentiated cells in culture for more than 10 passages. The isolated and cultured MSC comprised a single phenotypic population and displayed a fibroblast-like morphology. Simple method induced hMSC exhibiting a neuronal phenotype, with a positive expression of NSE, NF-M and nestin at 5 hours. But the neuron-like cells did not express the glial astrocyte marker GFAP. Conclusion It suggests that hMSC can be differentiated into neurons in vitro .

The study of expression of the basic fibroblast growth factor in normal and degenerated human intervertebral disc / 中华骨科杂志

Objective To investigate the expression of the basic fibroblast growth factor (bFGF) in normal and degenerated human intervertebral discs in order to determine whether bFGF is related to degeneration of the intervertebral disc. Methods The specimens of the intervertebral discs from 11 male and 19 female patients undergone lumbar disc herniation surgery (observation group) and 6 patients with scoliosis following anterior release (control group) were harvested. The tissues were histologically observed to confirm their degenerated or normal status and then conducted for the expression of bFGF and its mRNA with immunohistochemistry and in situ hybridization. Results In the observation group, all the samples were found to be degenerated disc tissue, and the positive rate of the expression of bFGF was 90% (27/30) with immunohistochemistry and 20% (6/30) with in situ hybridization. In the control group, all the samples were shown to be normal disc tissues, and had negative expression of bFGF with immunohistochemistry and in situ hybridization. The positive rate of the expression of bFGF showed significant difference between the two groups. Conclusion The expression of bFGF showed significant difference between the degenerated and normal intervertebral discs, which indicated that the bFGF might promote proliferation of chondrocyte and synthesis of extracellular matrix in the degenerated discs as a proliferation stimulating factor.

Effect of basic fibroblast growth factor on synthesis of C-type natriuretic peptide in cultured human umbilical vein endothelial cells / 中国病理生理杂志

AIM: To investigate the effect of basic fibroblast growth factor (bFGF) on C-type natriuretic peptide (CNP) production, release and mRNA expression. METHODS: Human endothelial cell cultured; CNP was measured by radioimmunoassay method;CNP mRNA expression was determined by RT-PCR technique. RESULTS: bFGF could augment CNP synthesis in human endothelial cells. Compared with control group,25 ng, 50 ng, 100 ng bFGF increased CNP contents in endothelial cells by 88% (P

Interleukin-6 induced upregulation of basic fibroblast growth factor isoform through phorbol ester-sensitive protein kinase C in rat vascular smooth muscle cells / 中国病理生理杂志

AIM: The role of protein kinase C(PKC) in the effect of Interleukin-6(IL-6) on basic fibroblast growth factor(bFGF) expression was investigated in rat vascular smooth muscle cells(VSMC). METHODS: Western-blotting was adopted to observe the variation of bFGF and its receptor type I isoforms expression. RESULTS: IL-6 increased all the three basic fibroblast growth factor isoforms in a dose-dependent (0-10.0 ?g/L) manner. The upregulatory activities peaked at 24 h as demonstrated . In addition, after exhaustion of intracellular phorbol ester-sensitive PKC, the upregulatory effects of IL-6 on bFGF exprssion in VSMC declined ( P

Metallothionein involvement in myocardial protection of basic fibroblast growth factor / 中国病理生理杂志

AIM: To observe whether metallothionein plays a role in cardiac protective effect of basic fibroblast growth factor on anoxic/reperfusion (A/R) injury in cultured cardiomyocytes and study the possible mechanism of cardiac protection by bFGF.METHODS: The present study made the model of myocyte A/R injury after having a 24 h incubation by bFGF(10 -10、10 -9、10 -8 mol/L) and bFGF(10 -9 mol/L)+PD 098059 respectively. We measured the levels of MT and MDA in myocytes, and the changes of LDH and protein in cultured medium. We also counted the number of viable cell in groups.RESULTS: The contents of myocardial MT were significantly increased after treatment by bFGF. The levels of MT in 10 -10 mol/L、10 -9 mol/L and 10 -8 mol/L bFGF treated groups increased 54%\, 62%\, 76% respectively, compared with the A/R group, and the number of viable cell were also greatly increased, LDH and protein leakage in cultured medium and MDA contents in myocyte were dramatically decreased in bFGF treated groups. All the protection were completely disappeared with the inhibition of MT production with PD 098059, the inhibitor of mitogen-activated protein kinase(MAPK).CONCLUSION: MT involves in the protection of bFGF in cultured cardiomyocytes. It might be related with activation of MAPKase.