mir-182-5p regulates all three phases of inflammation, proliferation, and remodeling during cutaneous wound healing.

Wound healing is a highly programmed process, in which any abnormalities result in scar formation. MicroRNAs are potent regulators affecting wound repair and scarification. However, the function of microRNAs in wound healing is not fully understood. Here, we analyzed the expression and function of microRNAs in patients with cutaneous wounds. Cutaneous wound biopsies from patients with either hypertrophic scarring or normal wound repair were collected during inflammation, proliferation, and remodeling phases. Fourteen candidate microRNAs were selected for expression analysis by qRT-PCR. The expression of genes involved in inflammation, angiogenesis, proliferation, and migration were measured using qRT-PCR. Cell cycle and scratch assays were used to explore the proliferation and migration rates. Flow cytometry analysis was employed to examine TGF-ß, αSMA and collagen-I expression. Target gene suggestion was performed using Enrichr tool. The results showed that miR-16-5p, miR-152-3p, miR-125b-5p, miR-34c-5p, and miR-182-5p were revealed to be differentially expressed between scarring and non-scarring wounds. Based on the expression patterns obtained, miR-182-5p was selected for functional studies. miR-182-5p induced RELA expression synergistically upon IL-6 induction in keratinocytes and promoted angiogenesis. miR-182-5p prevented keratinocyte migration, while overexpressed TGF-ß3 following induction of inflammation. Moreover, miR-182-5p enhanced fibroblast proliferation, migration, differentiation, and collagen-1 expression. FoxO1 and FoxO3 were found to potentially serve as putative gene targets of miR-182-5p. In conclusion, miR-182-5p is differentially expressed between scarring and non-scarring wounds and affect the behavior of cells involved in cutaneous wound healing. Deregulated expression of miR-182-5p adversely affects the proper transition of wound healing phases, resulting in scar formation.

Immunomodulatory performance of GMP-compliant, clinical-grade mesenchymal stromal cells from four different sources.

Inflammatory and autoimmune diseases are among the most challenging disorders for health care professionals that require systemic immune suppression which associates with various side effects. Mesenchymal stromal cells (MSCs) are capable of regulating immune responses, mainly through paracrine effects and cell-cell contact. Since MSCs are advanced therapy medicinal products (ATMPs), they must follow Good Manufacturing Practice (GMP) regulations to ensure their safety and efficacy. In this study, we evaluated the immunomodulatory effects of GMP-compliant clinical grade MSCs obtained from four different sources (bone marrow, adipose tissue, Wharton's Jelly, and decidua tissue) on allogeneic peripheral blood mononuclear cells (PBMCs). Our results revealed that WJ-MSCs were the most successful group in inhibiting PBMC proliferation as confirmed by BrdU analysis. Moreover, WJ-MSCs were the strongest group in enhancing the regulatory T cell population of PBMCs. WJ-MSCs also had the highest secretory profile of prostaglandin E2 (PGE-2), anti-inflammatory cytokine, while interleukin-10 (IL-10) secretion was highest in the DS-MSC group. DS-MSCs also had the lowest secretion of IL-12 and IL-17 inflammatory cytokines. Transcriptome analysis revealed that WJ-MSCs had the lowest expression of IL-6, while DS-MSCs were the most potent group in the expression of immunomodulatory factors such as hepatocyte growth factor (HGF) and transforming growth factor-ß (TGF- ß). Taken together, our results indicated that GMP-compliant Wharton's Jelly and decidua-derived MSCs showed the best immunomodulatory performance considering paracrine factors.

CRISPR/Cas9-Mediated Generation of COL7A1-Deficient Keratinocyte Model of Recessive Dystrophic Epidermolysis Bullosa.

OBJECTIVE: Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic skin fragility and ultimately lethal blistering disease caused by mutations in the COL7A1 gene which is responsible for coding type VII collagen. Investigating the pathological mechanisms and novel candidate therapies for RDEB could be fostered by new cellular models. The aim of this study was to employ CRISPR/Cas9 technology in the development of immortalized COL7A1-deficient keratinocyte cell lines intended for application as a cellular model for RDEB in ex vivo studies. MATERIALS AND METHODS: In this experimental study, we used transient transfection to express COL7A1 -targeting guide RNA (gRNA) and Cas9 in HEK001 immortalized keratinocyte cell line followed by enrichment with fluorescent-activated cell sorting (FACS) via GFP expressing cells (GFP+ HEK001). Homogenous single-cell clones were then isolated, genotyped, and evaluated for type VII collagen expression. We performed a scratch assay to confirm the functional effect of COL7A1 knockout. RESULTS: We achieved 46.1% (P<0.001) efficiency of in/del induction in the enriched transfected cell population. Except for 4% of single nucleotide insertions, the remaining in/dels were deletions of different sizes. Out of nine single expanded clones, two homozygous and two heterozygous COL7A1-deficient cell lines were obtained with defined mutation sequences. No off-target effect was detected in the knockout cell lines. Immunostaining and western blot analysis showed lack of type VII collagen (COL7A1) protein expression in these cell lines. We also showed that COL7A1-deficient cells had higher motility compared to their wild-type counterparts. CONCLUSION: We reported the first isogenic immortalized COL7A1-deficient keratinocyte lines that provide a useful cell culture model to investigate aspects of RDEB biology and potential therapeutic options.

Assessment of the Efficacy of an LL-37-Encapsulated Keratin Hydrogel for the Treatment of Full-Thickness Wounds.

Wound healing remains a burdensome healthcare problem due to moisture loss and bacterial infection. Advanced hydrogel dressings can help to resolve these issues by assisting and accelerating regenerative processes such as cell migration and angiogenesis because of the similarities between their composition and structure with natural skin. In this study, we aimed to develop a keratin-based hydrogel dressing and investigate the impact of the delivery of LL-37 antimicrobial peptide using this hydrogel in treating full-thickness rat wounds. Therefore, oxidized (keratose) and reduced (kerateine) keratins were utilized to prepare 10% (w/v) hydrogels with different ratios of keratose and kerateine. The mechanical properties of these hydrogels with compressive modulus of 6-32 kPa and tan δ <1 render them suitable for wound healing applications. Also, sustained release of LL-37 from the keratin hydrogel was achieved, which can lead to superior wound healing. In vitro studies confirmed that LL-37 containing 25:75% of keratose/kerateine (L-KO25:KN75) would result in significant fibroblast proliferation (∼85% on day 7), adhesion (∼90 cells/HPF), and migration (73% scratch closure after 12 h and complete closure after 24 h). Also, L-KO25:KN75 is capable of eradicating both Gram-negative and Gram-positive bacteria after 18 h. According to in vivo assessment of L-KO25:KN75, wound closure at day 21 was >98% and microvessel density (>30 vessels/HPF at day 14) was significantly superior in comparison to other treatment groups. The mRNA expression of VEGF and IL-6 was also increased in the L-KO25:KN75-treated group and contributed to proper wound healing. Therefore, the LL-37-containing keratin hydrogel ameliorated wound closure, and also angiogenesis was enhanced as a result of LL-37 delivery. These results suggested that the L-KO25:KN75 hydrogel could be a sustainable substitute for skin tissue regeneration in medical applications.

Comparison of Skin Transcriptome between Responder and Non-Responder Vitiligo Lesions to Cell Transplantation: A Clinical Trial Study.

Objective: Autologous transplantation of epidermal cells has been used increasingly to treat vitiligo patients and is a simple, safe, and relatively efficient method. However, the outcome is not always satisfactory, and some patients show less or no response to this treatment. This study was evaluated to identify genes expressed differently among responders and non-responders to cell transplantation to find potential markers that could predict 'patients' responses to this type of cell therapy. Materials and Methods: Eleven stable vitiligo patients who received autologous epidermal cell transplantation were included in this clinical trial study. Before cell transplantation, skin samples were obtained from the recipient's vitiligo lesions. After epidermal cell transplantation, patients were followed for at least six months to assess the response to epidermal cell injection. RNA sequencing was used to determine potential gene expression profile differences between responder and non-responder vitiligo patients. Results: The RNA sequencing results showed differences in expression levels of 470 genes between the skin specimens of responder versus non-responder patients. There were 269 up-regulated genes and 201 down-regulated genes. Upregulated genes were involved in processes, such as Fatty Acid Omega Oxidation. Down-regulated genes were related to PPAR signaling pathway, and estrogen signaling pathway. Among the most differentially expressed genes (DEGs) with the most altered RNA expression levels in responders versus non-responder patients, we selected three genes (up-regulated genes KRTAP10-11 and down-regulated genes IP6K2 and C9) as potential biomarkers, which are involved in associated pathways. Conclusion: Based on our findings, it is estimated that proposed genes might predict the response of vitiligo patients to cell therapy. However, further studies are required to clarify the role of these genes in pathogenesis and to characterize gene expression in a larger number of vitiligo patients in the context of epidermal cell transplantation therapy (registration number: IRCT201508201031N16).

The Emerging Therapeutic Targets for Scar Management: Genetic and Epigenetic Landscapes.

BACKGROUND: Wound healing is a complex process including hemostasis, inflammation, proliferation, and remodeling during which an orchestrated array of biological and molecular events occurs to promote skin regeneration. Abnormalities in each step of the wound healing process lead to reparative rather than regenerative responses, thereby driving the formation of cutaneous scar. Patients suffering from scars represent serious health problems such as contractures, functional and esthetic concerns as well as painful, thick, and itchy complications, which generally decrease the quality of life and impose high medical costs. Therefore, therapies reducing cutaneous scarring are necessary to improve patients' rehabilitation. SUMMARY: Current approaches to remove scars, including surgical and nonsurgical methods, are not efficient enough, which is in principle due to our limited knowledge about underlying mechanisms of pathological as well as the physiological wound healing process. Thus, therapeutic interventions focused on basic science including genetic and epigenetic knowledge are recently taken into consideration as promising approaches for scar management since they have the potential to provide targeted therapies and improve the conventional treatments as well as present opportunities for combination therapy. In this review, we highlight the recent advances in skin regenerative medicine through genetic and epigenetic approaches to achieve novel insights for the development of safe, efficient, and reproducible therapies and discuss promising approaches for scar management. KEY MESSAGE: Genetic and epigenetic regulatory switches are promising targets for scar management, provided the associated challenges are to be addressed.

Validation and diagnostic accuracy of coin rotation task for manual dexterity and coordination in children with specific learning disorder.

BACKGROUND: This study aimed at determining validity, reliability, and diagnostic accuracy of Coin Rotation Task (CRT) in assessing manual dexterity and coordination of children with specific learning disorder (SLD). METHODS: In this non-experimental cross-sectional study, 120 children (typically developing children = 60, children with SLD = 60, mean age ± SD =9.18 ± 0.55) were recruited. Test-retest reliability and construct validity of CRT were assessed. Multivariate regression analysis was performed on CRT scores by considering age and gender as covariates and children with SLD with mild dexterity impairment and severe dexterity impairment (SDI) as outcome variables. Receiver-operating characteristics (ROC) curve analysis was carried out to derive validity parameters. RESULTS: Test-retest reliability of the CRT scores in both subtests were excellent in children with SLD (ICC2,1: 0.93-0.95) and good to excellent in typically developing children (ICC2,1: 0.72-0.82). Acceptable construct validity of CRT was also found. The CRT cut-off points were 23 (sensitivity= 89.29%, specificity= 70.37%) and 28 (sensitivity= 80.33%, specificity= 86.36%) for discriminating children with SLD and SDI from typically developing children in dominant and non-dominant hand, respectively. CONCLUSIONS: The present study indicated good to excellent test-retest reliability, acceptable validity, and high diagnostic accuracy for diagnosing children with SLD based on their dexterity impairment level.Implications for RehabilitationThe Coin Rotation Task (CRT) was modified and validated for use in children.The CRT is a reliable and valid tool with high diagnostic accuracy.The CRT has a good ability for discriminating children with specific learning disorder with severe dexterity impairment form typically developing children.Treatment plans and research designs can be performed by using this valid, reliable, and easy to administer tool.

Defining microRNA signatures of hair follicular stem and progenitor cells in healthy and androgenic alopecia patients.

BACKGROUND: The exact pathogenic mechanism causes hair miniaturization during androgenic alopecia (AGA) has not been delineated. Recent evidence has shown a role for non-coding regulatory RNAs, such as microRNAs (miRNAs), in skin and hair disease. There is no reported information about the role of miRNAs in hair epithelial cells of AGA. OBJECTIVES: To investigate the roles of miRNAs affecting AGA in normal and patient's epithelial hair cells. METHODS: Normal follicular stem and progenitor cells, as well as follicular patient's stem cells, were sorted from hair follicles, and a miRNA q-PCR profiling to compare the expression of 748 miRNA (miRs) in sorted cells were performed. Further, we examined the putative functional implication of the most differentially regulated miRNA (miR-324-3p) in differentiation, proliferation and migration of cultured keratinocytes by qRT-PCR, immunofluorescence, and scratch assay. To explore the mechanisms underlying the effects of miR-324-3p, we used specific chemical inhibitors targeting pathways influenced by miR-324-3p. RESULT: We provide a comprehensive assessment of the "miRNome" of normal and AGA follicular stem and progenitor cells. Differentially regulated miRNA signatures highlight several miRNA candidates including miRNA-324-3p as mis regulated in patient's stem cells. We find that miR-324-3p promotes differentiation and migration of cultured keratinocytes likely through the regulation of mitogen-activated protein kinase (MAPK) and transforming growth factor (TGF)-ß signaling. Importantly, pharmacological inhibition of the TGF-ß signaling pathway using Alk5i promotes hair shaft elongation in an organ-culture system. CONCLUSION: Together, we offer a platform for understanding miRNA dynamic regulation in follicular stem and progenitor cells in baldness and highlight miR-324-3p as a promising target for its treatment.

Long-Term Follow-up of Autologous Fibroblast Transplantation for Facial Contour Deformities, A Non-Randomized Phase IIa Clinical Trial.

OBJECTIVE: Recently, the promising potential of fibroblast transplantation has become a novel modality for skin rejuvenation. We investigated the long-term safety and efficacy of autologous fibroblast transplantation for participants with mild to severe facial contour deformities. MATERIALS AND METHODS: In this open-label, single-arm phase IIa clinical trial, a total of 57 participants with wrinkles (n=37, 132 treatment sites) or acne scars (n=20, 36 treatment sites) who had an evaluator's assessment score of at least 2 out 7 (based on a standard photo-guide scoring) received 3 injections of autologous cultured fibroblasts administered at 4-6 week intervals. Efficacy evaluations were performed at 2, 6, 12, and 24 months after the final injection based on evaluator and patient's assessment scores. RESULTS: Our study showed a mean improvement of 2 scores in the wrinkle and acne scar treatment sites. At sixth months after transplantation, 90.1% of the wrinkle sites and 86.1% of the acne scar sites showed at least a one grade improvement on evaluator assessments. We also observed at least a 2-grade improvement in 56.1% of the wrinkle sites and 63.9% of the acne scar sites. A total of 70.5% of wrinkle sites and 72.2% of acne scar sites were scored as good or excellent on patient assessments. The efficacy outcomes remained stable up to 24-month. We did not observe any serious adverse events during the study. CONCLUSION: These results have shown that autologous fibroblast transplantation could be a promising remodeling modality with long-term corrective ability and minimal adverse events (Registration Number: NCT01115634).

Copper nanoparticles promote rapid wound healing in acute full thickness defect via acceleration of skin cell migration, proliferation, and neovascularization.

Worldwide, impaired wound healing leads to a large burden of morbidity and mortality. Current treatments have several limitations. Recently, nanomaterials such as copper nanoparticles (CuNPs) have attracted considerable research interest. Here, we investigated the potential therapeutic effect of various CuNPs concentrations (1 µM, 10 µM, 100 µM, 1 mM, and 10 mM) and sizes (20 nm, 40 nm, 80 nm) in wound healing. Our results revealed that the 10 µM concentration of 40 nm CuNPs and the 1 µM concentration of 80 nm CuNPs were not toxic to the cultured fibroblast, endothelial, and keratinocyte cells, and also 1 µM concentration of 80 nm CuNPs enhanced endothelial cell migration and proliferation. Extensive assessment of in vivo wound healing demonstrated that the 1 µM concentration of 80 nm CuNPs accelerated wound healing over a shorter time via formation of granulation tissue and higher new blood vessels. Importantly, serum biochemical analysis confirmed that the 40 nm CuNP (10 µM) and 80 nm CuNP (1 µM) did not show any accumulation in the liver during wound healing. Overall, our results have indicated that the 1 µM concentration of 80 nm CuNPs is a promising NP for wound healing applications without adverse side effects.

Contribution of NOTCH signaling pathway along with TNF-α in the intestinal inflammation of ulcerative colitis.

AIM: The aim of this study was to determine gene expression levels of TNF-α, NOTCH1, and HES1 in patients with UC. BACKGROUND: Intestinal inflammation and epithelial injury are the leading actors of inflammatory bowel disease (IBD), causing an excessive expression of pro-inflammatory cytokines such as TNF-α. Also, target genes of NOTCH signaling are involved in the regulation of intestinal homeostasis. Previous studies have demonstrated that TNF-α increases in ulcerative colitis (UC) patients, but the relationship between TNF-α and NOTCH signaling pathway in UC etiopathology needs further study. METHODS: Twelve active UC patients and twelve healthy controls were enrolled in this study. RNA was extracted and the mRNA expression levels of TNF-α, NOTCH1, and HES1 were examined using real-time PCR analyses. Further, transcriptome data deposited in Gene Expression Omnibus (GEO) database were analyzed to detect the differential expression of TNF superfamily and NOTCH1 gene in IBD patients. Finally, the interaction of TNF-α and NOTCH signaling was obtained from The SIGnaling Network Open Resource 2.0 (SIGNOR 2.0) database. RESULTS: The transcription levels of TNF-α, NOTCH1, and HES1 genes were significantly elevated in UC patients compared with control (p < 0.05). In addition, GEO results confirmed our expression results. SIGNOR analysis showed that TNF-α interacts with NOTCH signaling components. CONCLUSION: Based on our data, we observed that NOTCH1 and HES1 in co-operation of TNF-α, may play an important role in pathogenesis of UC. The members of NOTCH signaling pathway can be ideal candidates to target the therapy of IBD.

Amniotic Membrane Extract Eye Drop Promotes Limbal Stem Cell Proliferation and Corneal Epithelium Healing.

OBJECTIVE: Human amniotic membrane (HAM) is used as a supporter for limbal stem cell (LSC) expansion and corneal surgery. The aim of study is to use HAM extracts from healthy donors to enhance proliferation of LSCs in vitro and in vivo. MATERIALS AND METHODS: In this interventional experimental study, the effective and cytotoxic doses of the amniotic membrane extract eye drops (AMEED) was assessed by adding different concentrations of AMEED (0-2.0 mg/ml) to LSC cultures for 14 days. Subsequently, the expression levels of ATP-binding cassette sub-family G member 2 (ABCG2, a putative stem cell marker), cytokeratin 3 (K3, corneal maker), K12 and K19 (corneal-conjunctival cell makers) were assessed by real-time polymerase chain reaction (PCR). In the second step, the corneal epithelium of 10 rabbits was mechanically removed, and the right eye of each rabbit was treated with 1 mg/ml AMEED [every 2 hours (group 1) or every 6 hours (group 2)]. The left eyes only received an antibiotic. The corneal healing process, conjunctival infection, degree of eyelid oedema, degree of photophobia, and discharge scores were evaluated during daily assessments. Finally, corneal tissues were biopsied for pathologic evidences. RESULTS: In comparison to the positive control [10% foetal bovine serum (FBS)], 0.1-1 mg/ml AMEED induced LSC proliferation, upregulated ABCG2, and downregulated K3. There were no remarkable differences in the expression levels of K12 and K19 (P>0.05). Interestingly, in the rabbits treated with AMEED, the epithelium healing duration decreased from 4 days in the control group to 3 days in the two AMEED groups, with lower mean degrees of eyelid oedema, chemosis, and infection compared to the control group. No pathologic abnormalities were observed in either of the AMEED groups. CONCLUSION: AMEED increases LSCs proliferation ex vivo and accelerates corneal epithelium healing in vivo without any adverse effects. It could be used as a supplement for LSC expansion in cell therapy.

A single-arm open-label clinical trial of autologous epidermal cell transplantation for stable vitiligo: A 30-month follow-up.

BACKGROUND: Recently, we introduced intralesional injection of autologous epidermal cells as a safe and feasible approach for transplantation in patients with stable vitiligo. This approach resulted in less pain during and after the procedure, no scarring or cobblestone formation at the recipient site, and was more feasible to perform on curved surfaces such as joints, lips, eyelids, ears, and face. OBJECTIVE: In this study, we aimed to investigate the long-term efficacy and safety of this transplantation technique. METHODS: In this open-label and single-arm clinical trial, we enrolled 300 patients with stable vitiligo. We obtained a partial thickness normo-pigmented skin specimen from the patients' thigh-buttock junction with an area of one tenth to one third of the recipient site area. The epidermal cell suspension was prepared by processing the autologous skin specimen. We injected the cell suspension into 1060 vitiligo patches in 300 patients. Patients did not use any adjuvant phototherapy during the study. An experienced dermatologist and patients respectively defined the repigmentation score and self-assessment score at regular follow-up visits for up to 30 months after treatment. The scores represented the repigmentation percentage as follows: 0 (0), I (1%-24%), II (25%-49%), III (50%-74%), and IV (75%-100%). RESULTS: The mean repigmentation score at 3 months post-transplantation was 1.12±0.73. A significant upward trend existed in the mean repigmentation score until 9 months after cell transplantation, when the mean repigmentation score reached to 1.98±1.20. At 9 months after treatment, repigmentation of >50% was obtained in 32.2% of treated patches. Acquired repigmentation remained stable in 79.3% of treated patches during the follow-up period. The number of received cells per cm2 positively influenced the repigmentation score. Patches located on face, neck and trunk showed significantly higher response to the treatment. CONCLUSION: The results of our study demonstrated efficacy and safety of autologus epidermal cell transplantation on repigmentation of vitiligo patches. The achieved repigmentation was stable in the majority of treated patches during the follow-up period.

Hair Follicle Generation by Injections of Adult Human Follicular Epithelial and Dermal Papilla Cells into Nude Mice.

OBJECTIVE: Dermal papilla and hair epithelial stem cells regulate hair formation and the growth cycle. Damage to or loss of these cells can cause hair loss. Although several studies claim to reconstitute hairs using rodent cells in an animal model, additional research is needed to develop a stable human hair follicle reconstitution protocol. In this study, we have evaluated hair induction by injecting adult cultured human dermal papilla cells and a mixture of hair epithelial and dermal papilla cells in a mouse model. MATERIALS AND METHODS: In this experimental study, discarded human scalp skins were used to obtain dermal papilla and hair epithelial cells. After separation, cells were cultured and assessed for their characteristics. We randomly allocated 15 C57BL/6 nude mice into three groups that received injections in their dorsal skin. The first group received cultured dermal papilla cells, the second group received a mixture of cultured epithelial and dermal papilla cells, and the third group (control) received a placebo [phosphate-buffered saline (PBS-)]. RESULTS: Histopathologic examination of the injection sites showed evidence of hair growth in samples that received cells compared with the control group. However, the group that received epithelial and dermal papilla cells had visible evidence of hair growth. PKH tracing confirmed the presence of transplanted cells in the new hair. CONCLUSION: Our data showed that injection of a combination of adult human cultured dermal papilla and epithelial cells could induce hair growth in nude mice. This study emphasized that the combination of human adult cultured dermal papilla and epithelial cells could induce new hair in nude mice.

Human Hair Reconstruction: Close, But Yet So Far.

Billions of dollars are annually invested in pharmaceutical industry and cosmetic sector with intent to develop new drugs and treatment strategies for alopecia. Because the hair looks an important characteristic of humans-an effective appendage in perception, expression of beauty, and preservation of self-esteem-the global market for hair loss treatment products is exponentially increasing. However, current methods to treat hair loss endure yet multiple challenges, such as unfavorable outcomes, nonpermanent and patient-dependent results, as well as unpredictable impacts, which limit their application. Over recent years, remarkable advances in the fields of regenerative medicine and hair tissue engineering have raised new hopes for introducing novel cell-based approaches to treat hair loss. Through cell-based approaches, it is possible to produce hair-like structures in the laboratory setting or manipulate cells in their native niche (in vivo lineage reprogramming) to reconstruct the hair follicle. However, challenging issues still exist with the functionality of cultured human hair cells, the proper selection of nonhair cell sources in cases of shortage of donor hair, and the development of defined culture conditions. Moreover, in the case of in vivo lineage reprogramming, selecting appropriate induction factors and their efficient delivery to guide resident cells into a hair fate-with the aim of reconstructing functional hair-still needs further explorations. In this study, we highlight recent advances and current challenges in hair loss treatment using cell-based approaches and provide novel insights for crucial steps, which must be taken into account to develop reproducible, safe, and efficient cell-based treatment.

Facile fabrication of egg white macroporous sponges for tissue regeneration.

The availability of 3D sponges combining proper biochemical, biophysical, and biomechanical properties with enhanced capacity of in vivo engraftment and vascularization is crucial in regenerative medicine. A simple process is developed to generate macroporous scaffolds with a well-defined architecture of interconnected pores from chicken egg white (EW), a material with protein- and growth factor-binding features which has not yet been employed in regenerative medicine. The physicomechanical properties and degradation rates of the scaffold are finely tuned by using varying concentrations of the cross-linker, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride, without alteration of the biochemical traits. In vitro, EW scaffolds supported active metabolism, proliferation, and migration of human dermal fibroblasts, thereby generating uniform cellular constructs. In vivo, subcutaneous implantation in mice reveals negligible immune reaction and efficient cell and tissue ingrowth. Angiogenesis into EW scaffolds is enhanced as compared to standard collagen type I sponges used as reference material, likely due to significantly higher adsorption of the proangiogenic factor vascular endothelial growth factor. In summary, a material is presented derived by facile processing of a highly abundant natural product. Due to the efficient subcutaneous engraftment capacity, the sponges can find utilization for soft tissue regeneration.

Treatment of Hypertrophic Scar in Human with Autologous Transplantation of Cultured Keratinocytes and Fibroblasts along with Fibrin Glue.

OBJECTIVE: Hypertrophic scar involves excessive amounts of collagen in dermal layer and may be painful. Nowadays, we can't be sure about effectiveness of procedure for hypertrophic scar management. The application of stem cells with natural scaffold has been the best option for treatment of burn wounds and skin defect, in recent decades. Fibrin glue (FG) was among the first of the natural biomaterials applied to enhance skin deformity in burn patients. This study aimed to identify an efficient, minimally invasive and economical transplantation procedure using novel FG from human cord blood for treatment of hypertrophic scar and regulation collagen synthesis. MATERIALS AND METHODS: In this case series study, eight patients were selected with hypertrophic scar due to full-thickness burns. Human keratinocytes and fibroblasts derived from adult skin donors were isolated and cultured. They were tested for the expression of cytokeratin 14 and vimentin using immunocytochemistry. FG was prepared from pooled cord blood. Hypertrophic scars were extensively excised then grafted by simply placing the sheet of FG containing autologous fibroblast and keratinocytes. Histological analyses were performed using Hematoxylin and eosin (H&E) and Masson's Trichrome (MT) staining of the biopsies after 8 weeks. RESULTS: Cultured keratinocytes showed a high level of cytokeratin 14 expression and also fibroblasts showed a high level of vimentin. Histological analyses of skin biopsies after 8 weeks of transplantation revealed re-epithelialization with reduction of hypertrophic scars in 2 patients. CONCLUSION: These results suggest may be the use of FG from cord blood, which is not more efficient than previous biological transporters and increasing hypertrophic scar relapse, but could lead to decrease pain rate.

Effect of vitamin D3 supplementation on TNF-α serum level and disease activity index in Iranian IBD patients.

AIM: The aim of the study was to assess the effectiveness of vitamin D3 [1, 25(OH)2D3] treatment in IBD with regard to tumor necrosis factor-alpha (TNF-α) serum level and clinical disease activity index (CDAI). BACKGROUND: Vitamin D has immune-regulatory functions in experimental inflammatory bowel disease (IBD) and vitamin D deficiency is common in IBD patients. PATIENTS AND METHODS: This was a randomized clinical trial on 108 IBD patients with serum 25-OHD levels less than 30ng/ml, which divided into vitamin D and control groups. Vitamin D group received 50000 IU vitamin D3 for 12 weeks. Before and after the study, TNF-α and 25-OHD serum levels were measured by ELISA method. Data were analyzed using paired t-test, chi-square test and Spearman correlation coefficient. P-values ​​less than 0.05 were considered statistically significant. RESULTS: Before the intervention no significant difference was found between baseline characteristics and TNF-α serum level of two groups. After intervention TNF-α serum level reduced but this reduction was not statistically significant (p= 0.07, 95% CI: -0.45 to 8.14). The mean serum 25-OHD level of vitamin D increased from 15.54 to 67.89, which was statistically significant (p= 0.00, 95% CI: -61.40 to -43.30). TNF-α level was also associated significantly with CDAI before (Spearman's rho: 0.3, p<0.0001) and after (Spearman's rho: 0.27, P=0.01) intervention. CONCLUSION: Oral supplementation vitamin D3 significantly increased serum vitamin D levels and insignificantly reduced serum TNF-α level. More studies with larger samples would be beneficial to assess vitamin D3 supplementation efficient effect in IBD.

Prevalence of hepatitis D virus in hepatitis B virus infected patients referred to Taleghani hospital, Tehran, Iran.

AIM: The aim of this study was to determine the prevalence of HDV infection between HBV chronic patients referred to gastroenterology ward of Taleghani hospital Tehran, Iran and also investigating the risk factors in acquiring the HDV infection. BACKGROUND: Hepatitis B virus (HBV) and Hepatitis D virus (HDV) are major public health issues. Worldwide there are approximately 350 million individuals chronically infected with the HBV. A significant part of them, including 15 to 20 million coinfected with HDV. Hepatitis Delta virus is transferred mostly through blood and body fluids. PATIENTS AND METHODS: HBV and HDV infections were evaluated by Enzyme-linked immunosorbent assay (ELISA). Liver functional tests were assessed through auto analyzer. Patients were interviewed and data along the test results were entered into SPSS program. We used chi-square, independent t-test and logistic regression for statistical analysis. RESULTS: 278 (54.6%) patients of the study group were male and 231 (45.4%) were female and the mean age of patients was 40.03 ± 14.93. From 509 patients, 39(7.7%) had anti-HDV antibody. In a uni-variable analysis, age (p=0.001), periodontal procedures (p=0.015), endoscopy (p=0.024) and colonoscopy (p=0.012) were significantly related to HDV seropositivity. After adjustment by logistic regression, age remained the only significant factor in acquiring HDV infection. CONCLUSION: We highly recommend the health care workers to strictly follow the disinfection protocols of medical instruments. Since HDV seroprevalence changes over time, regular epidemiological studies are necessary to monitor the epidemiological trend of infection.

Low frequency of human T-cell lymphotropic virus 1 antibodies in Iranian gastric cancer patients in comparison to controls.

There is some evidence that human T-cell lymphotropic virus (HTLV-1) infection has a reverse association with gastric cancer (GC). Data about this association in the Iranian population are scarce. In this study we therefore assessed the frequency of anti-HTLV-1 antibody in GC patients and compare it to antibody presence in healthy individuals in Iranian population. This case control study was performed between 2008-2011 on 201 GC patients and 219 control subjects. HTLV-1 antibodies were assessed by ELISA and the positive results were confirmed by western blotting. Totals of 201 gastric cancer patients and 219 controls were enrolled in this study. The tumors in the majority of patients (45.3%) were in the distal (non-cardia) area. Mean age of patients at the time of diagnosis was 59.2±12.5 and mean age of controls was 57.7±11.3. While only one GC patient (0.5%) was positive for HTLV-1 antibody, there were four individuals (1.89%) from the control group with antibodies. In addition, smoking had statistically significant relationship with cancer (P=0.001). Our study showed that the frequency of HTLV-1 antibody in patients was lower than in controls, similar to the results obtained in Japan. Further investigations with a larger sample size are needed in order to determine the association between GC and HTLV-1 infection in Iran.