Co-axial electrospinning of PLLA shell, collagen core nanofibers for skin tissue engineering.

The main functions of wound dressing biomaterials are to promote a moist environment in the wound while protecting the area from mechanical injury and microbial contamination. Furthermore, the scaffold used for skin tissue engineering must mimic epidermal and dermal layers as well as support the growth of keratinocytes and fibroblasts. In this study, PLLA (shell) and EGF-encapsulated collagen (core) nanofibers were produced by coaxial electrospinning at 25 kV potential, 12 cm collector distance, and 0.125 mL/h flow rate of PLLA (15%) and collagen (4%). The bilayer structure was produced by gelling GeIMA in between two nanofiber membranes to imitate the epidermal and dermal layers of skin. Cytocompatibility properties of nanofiber membrane and bilayer structure were characterized by mono- and coculture of keratinocytes (HaCaT) and fibroblasts (3T3), respectively. TEM revealed that the PLLA shell and collagen core thicknesses were about 60 and 115 nm, respectively. Oxygen and water vapor could pass through the GeIMA- integrated bilayer nanofiber membranes. The presence of EGF in nanofibers could increase cell proliferation. Fluorescence and SEM imaging showed that HaCaT and 3T3 could cover the membrane after 14 days of monoculture. Cocultures showed a reduction in the proliferation of cells in the first week and a recovery during the second and third weeks. In a mechanical bioreactor, cocultured bilayer membranes formed interlocked polygonal keratinocyte cells. These results showed that the bilayer nanofiber membrane and GeIMA combination provided cell compatibility. Furthermore, the use of a mechanical reactor was found to be effective in the formation of a functional keratinocyte layer by stimulating cells.

The utility of TNF-alfa, IL-6 and IL-10 in the diagnosis and/or follow-up food allergy

Background: Several pro-inflammatory and anti-inflammatory mediators play a role in the immunopathogenesis of food allergy (FA). The aim of this study was to investigate the utility of serum biomarkers like interleukin (IL)-10, TNF-alfa, and IL-6 in the diagnosis and/or follow-up of FA. Methods: Sixty (25 females, 41.6%) newly diagnosed FA patients [IgE mediated (group-1, n = 37), non-IgE (group-2, n = 23)] with a median age of nine (1-33) months were enrolled. Twenty-four healthy children with a median age of eight (1-36) months constituted the control group (CG). In all the subjects, serum TNF-alfa, IL-6 and IL-10 levels were evaluated at the time of diagnosis and reassessed four weeks after therapeutic elimination diet (TED). Results: The mean white blood cell count and median absolute eosinophile count of the CG were significantly lower than group-1 (p values were 0.019 and 0.006, respectively). The mean absolute neutrophile count and the median IL-6 were significantly higher in group-1 when compared with group-2 (p values were 0.005 and 0.032, respectively. Median TNF-alfa and IL-6 levels were significantly higher in the pre-TED among all patients (p values were 0.005 and 0.018, respectively). In group-1, median TNF-α and IL-6 levels decreased significantly after TED (p values were 0.01 and 0.029, respectively). Conclusions: Our findings support the role of inflammation in the pathogenesis of FA. Serum TNF-alfa and IL-6 levels may be useful markers for follow-up in FA, especially among IgE-mediated FA patients. Evaluation of IL-10 results was not sufficient for an interpretation of clinical tolerance

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The utility of TNF-α, IL-6 and IL-10 in the diagnosis and/or follow-up food allergy.

BACKGROUND: Several pro-inflammatory and anti-inflammatory mediators play a role in the immunopathogenesis of food allergy (FA). The aim of this study was to investigate the utility of serum biomarkers like interleukin (IL)-10, TNF-α, and IL-6 in the diagnosis and/or follow-up of FA. METHODS: Sixty (25 females, 41.6%) newly diagnosed FA patients [IgE mediated (group-1, n=37), non-IgE (group-2, n=23)] with a median age of nine (1-33) months were enrolled. Twenty-four healthy children with a median age of eight (1-36) months constituted the control group (CG). In all the subjects, serum TNF-α, IL-6 and IL-10 levels were evaluated at the time of diagnosis and reassessed four weeks after therapeutic elimination diet (TED). RESULTS: The mean white blood cell count and median absolute eosinophile count of the CG were significantly lower than group-1 (p values were 0.019 and 0.006, respectively). The mean absolute neutrophile count and the median IL-6 were significantly higher in group-1 when compared with group-2 (p values were 0.005 and 0.032, respectively. Median TNF-α and IL-6 levels were significantly higher in the pre-TED among all patients (p values were 0.005 and 0.018, respectively). In group-1, median TNF-α and IL-6 levels decreased significantly after TED (p values were 0.01 and 0.029, respectively). CONCLUSIONS: Our findings support the role of inflammation in the pathogenesis of FA. Serum TNF-α and IL-6 levels may be useful markers for follow-up in FA, especially among IgE-mediated FA patients. Evaluation of IL-10 results was not sufficient for an interpretation of clinical tolerance.

The frequency of celiac disease in children with autoimmune thyroiditis.

BACKGROUND AND AIMS: Although the presence of autoimmune thyroiditis (AT) in celiac disease (CD) has been well documented among adults, CD in AT has been less reported in children. We aimed to investigate the frequency of CD in children with AT. Materials and Methods : This prospective study was carried out from October 2015 to August 2016 and included 66 patients with AT. Firstly, total IgA and tissue transglutaminase antibody (tTG) IgA levels were measured. Those with increased level of tTG IgA were tested for anti-endomysium IgA antibodies (EMA). Patients with positive EMA underwent gastroduodenoscopy for a definitive diagnosis of CD. RESULTS: Sixty-six patients with AT (52 female) with mean age of 14.68 ± 3.18 years were enrolled. IgA deficiency was found in four patients. Only three of 66 patients (4.5%) were positive for tTG IgA. Patients positive for tTG IgA were then tested for EMA, and only one of them (1.5%) had positive EMA antibodies. Gastroduodenoscopy was performed in this patient. The result of pathological investigation was compatible with CD. Furthermore, one patient with AT had been diagnosed with CD previously. CONCLUSIONS: Two (3.0%) of 66 patients with AT were found to have CD. According to the results, we assume that there is a close relationship between CD and AT disease. However, there is a need for multicentric, prospective studies that would support our findings.

Construction of a patterned hydrogel-fibrous mat bilayer structure to mimic choroid and Bruch's membrane layers of retina.

Deterioration of retina and death of the retinal cells due to age, diabetes, or occlusion can cause retinal degeneration which leads to loss of vision. In this study, it is aimed to design a bilayered matrix to mimic the choroid and the Bruch's membrane of the retinal tissue. As choroid, a microchanneled network resembling a fractal tree design was fabricated by photolithography over photo-cross-linkable methacrylated hyaluronic acid hydrogel. Gelatin or collagen was immobilized into the microchannels to enhance adherence of Human Umbilical Vein Endothelial Cells (HUVEC). At late culture periods (2 weeks), formation of tubular structures due to proliferation of the attached cells was observed. As Bruch's membrane, an electrospun fibroin nanofiber mat was produced to grow retinal pigment epithelium (RPE) cells on. Cellular interactions between RPE and HUVEC in the microchannels were investigated in a coculture model in a noncontact mode. It was deduced that by combining the RPE layer on the highly permeable Bruch's membrane with the choroid layer populated by HUVECs, a retinal substitute which might have a potential for use in the treatment of retinal diseases is possible. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2166-2177, 2016.

CITRIN DEFICIENCY: AN INFANT INCIDENTALLY DETECTED BY PHENYLKETONURIA SCREENING WITH A NOVEL MUTATION IN SLC25A13 GENE.

We report the first Turkish patient with citrin deficiency detected incidentally by phenylketonuria screening. Mild cholestasis, increased α-fetoprotein level, aminoacidemia including citrulline and coagulation disorder suggested citrin deficiency. Screening the SLC25A13 gene revealed compound heterozygosity harboring a novel mutation, c.851-854delGTAT (p.M285Pfs*2)/ p.I290T (c.869T>C). Progression to type II citrullinemia was considered due to hyperammonemia episodes resulting from high carbohydrate/low protein diet. High protein/low carbohydrate diet resulted in cessation of hyperammonemia episodes, reversal of hepatic dysfunction and steatohepatitis. Our report illustrates the importance of awareness on citrin deficiency.

Self-assembly in nature: using the principles of nature to create complex nanobiomaterials.

Self-assembly is a ubiquitous process in biology where it plays numerous important roles and underlies the formation of a wide variety of complex biological structures. Over the past two decades, materials scientists have aspired to exploit nature's assembly principles to create artificial materials, with hierarchical structures and tailored properties, for the fabrication of functional devices. Toward this goal, both biological and synthetic building blocks have been subject of extensive research in self-assembly. In fact, molecular self-assembly is becoming increasingly important for the fabrication of biomaterials because it offers a great platform for constructing materials with high level of precision and complexity, integrating order and dynamics, to achieve functions such as stimuli-responsiveness, adaptation, recognition, transport, and catalysis. The importance of peptide self-assembling building blocks has been recognized in the last years, as demonstrated by the literature available on the topic. The simple structure of peptides, as well as their facile synthesis, makes peptides an excellent family of structural units for the bottom-up fabrication of complex nanobiomaterials. Additionally, peptides offer a great diversity of biochemical (specificity, intrinsic bioactivity, biodegradability) and physical (small size, conformation) properties to form self-assembled structures with different molecular configurations. The motivation of this review is to provide an overview on the design principles for peptide self-assembly and to illustrate how these principles have been applied to manipulate their self-assembly across the scales. Applications of self-assembling peptides as nanobiomaterials, including carriers for drug delivery, hydrogels for cell culture and tissue repair are also described.

Microfluidic fabrication of self-assembled peptide-polysaccharide microcapsules as 3D environments for cell culture.

We report a mild cell encapsulation method based on self-assembly and microfluidics technology. Xanthan gum, an anionic polysaccharide, was used to trigger the self-assembly of a positively charged multidomain peptide. The self-assembly resulted in the formation of a nanofibrous matrix and using a microfluidic device, microcapsules with homogeneous size were fabricated. The properties and performance of xanthan-peptide microcapsules were optimized by changing peptide/polysaccharide ratio and their effects on the microcapsule permeability and mechanical stability were analyzed. The effect of microcapsule formulation on viability and proliferation of encapsulated chondrocytic (ATDC5) cells was also investigated. The encapsulated cells were metabolically active, showing an increased viability and proliferation over 21 days of in vitro culture, demonstrating the long-term stability of the self-assembled microcapsules and their ability to support and enhance the survival of encapsulated cells over a prolonged time. Self-assembling materials combined with microfluidics demonstrated to be an innovative approach in the fabrication of cytocompatible matrix for cell microencapsulation and delivery.

Encapsulation and survival of a chondrocyte cell line within xanthan gum derivative.

A chemical derivative of xanthan gum polysaccharide is investigated as a new artificial matrix for the encapsulation of chondrocytic cells. Toward this goal, a novel micro-droplet generator is developed to produce microcapsules. Microcapsules with an average diameter of 500 µm, smooth surface, and homogeneous size distribution are obtained. ATDC5 cells encapsulated in carboxymethyl xanthan (CMX) microcapsules remain viable and are observed to proliferate for prolonged culture periods with enhanced metabolic activity. Furthermore, retention of the chondrogenic phenotype is exhibited by the cells within CMX, suggesting the ability of this material to be applied in cell-delivery therapies.

Enzymatic degradation behavior and cytocompatibility of silk fibroin-starch-chitosan conjugate membranes.

The objective of this study was to investigate the influence of silk fibroin and oxidized starch conjugation on the enzymatic degradation behavior and the cytocompatability of chitosan based biomaterials. The tensile stress of conjugate membranes, which was at 50 Megapascal (MPa) for the lowest fibroin and starch composition (10 weight percent (wt.%)), was decreased significantly with the increased content of fibroin and starch. The weight loss of conjugates in α-amylase was more notable when the starch concentration was the highest at 30 wt.%. The conjugates were resistant to the degradation by protease and lysozyme except for the conjugates with the lowest starch concentration. After 10 days of cell culture, the proliferation of osteoblast-like cells (SaOS-2) was stimulated significantly by higher fibroin compositions and the DNA synthesis on the conjugate with the highest fibroin (30 wt.%) was about two times more compared to the native chitosan. The light microscopy and the image analysis results showed that the cell area and the lengths were decreased significantly with higher fibroin/chitosan ratio. The study proved that the conjugation of fibroin and starch with the chitosan based biomaterials by the use of non-toxic reductive alkylation crosslinking significantly improved the cytocompatibility and modulated the biodegradation, respectively.

Microchannel-patterned and heparin micro-contact-printed biodegradable composite membranes for tissue-engineering applications.

Microchannel-patterned starch-poly(capro-lactone)/hydydroxyapatite (SPCL-HA) and starch-poly(lactic acid) (SPLA) composite membranes were produced for use as a laminated tissue-engineering scaffold that incorporates both physical and biochemical patterns. For this purpose, SPCL (30% starch) blended with inorganic hydroxyl apatite (50%) and SPLA (50% starch) membranes were made with compressive moulding. Consequently, the microchannel structures (width 102 µm, 174 µm intervals) were developed on the composite membranes by means of micro-patterned metal mould(s) and hydraulic pressing. An elastomer poly(dimetylsiloxane) stamp was used to transfer heparin as a biochemical cue over the microchannel surfaces by micro-contact printing (µCP). Toluidine blue staining of developed capillaries and heparin µCP-coated membranes showed that heparin was transferred predominantly over the microchannel surfaces. Fibroblast cell culture over the microchannel-formed and heparin µCP-modified SPCL-HA and SPLA membranes showed distinct growth patterns. In contrast to the uniform cell layer formed on unmodified microchannels, the cells were bridging across the grooves of heparin-printed microchannels. At extended culture periods, the heparin-printed microchannels were covered with a layer of fibroblast cells without cellular ingrowths inside. This study indicated that the topographical pattern could induce an organization of fibroblasts only with the biochemical cue and the cells' functions can be controlled spatially over the microchannels by using both cues.

HLA-DQ alleles in patients with celiac disease in Turkey.

The prevalence of the HLA-DQA1 and DQB1 alleles in 55 Turkish children with celiac disease and 50 control subjects was investigated by using an allele-specific DNA-based polymerase chain reaction-sequence-specific primer (PCR-SSP) method. The frequency of the DQA1*0501 and DQB1*02 alleles was higher in celiac patients than in controls. The DQA1B1 (*0501; *02) haplotype was present in 46 (83.6%) patients and only in 12 (24%) controls. The remaining 9 celiac patients which were negative for DQA1B1 (*0501;*02) carried the DQA1B1 (*03;*0302) haplotype. We found an excess homozygosity of the DQB1*02 allele and the DQA1B1 (*0501;*02) haplotype in the patients. No statistically significant correlation was found between the homozygosity of this haplotype or the DQB1*02 allele and an earlier onset of the disease.

Human leukocyte antigens in Turkish pediatric celiac patients.

With the aim to determine the frequency of human leukocyte antigen phenotypes of celiac disease in Turkey, thirty celiac patients fulfilling the European Society of Pediatric Gastroenterology and Nutrition criteria were included in the study. The mean age of the study population was 5.8 +/- 4.3 years and of the control subjects was 32.6 +/- 6.7 years. The human leukocyte antigens -A, -B, -DR and -DQ were studied serologically by micro lymphocytotoxic reaction. It was found that human leukocyte antigens A-25(10), -B8, -DR18(3) and -DQ2 were more significantly frequent in the celiac population than in the control group. Children with antigen -B8 showed a five times higher risk for celiac disease and those with antigen -DQ2 showed a nine times higher risk. It was determined that human leukocyte antigen -B4 had a protective role in celiac disease. The study suggests that the human leukocyte antigen -A25(10) is a phenotype particularly encountered in Turkish pediatric celiac patients.

A case of vertical transmission of hepatitis A virus infection.

We present a case of hepatitis A infection in a 2.5-month-old male who became icteric after 18 d of birth. The diagnosis of hepatitis A was made by compatible clinical symptoms, laboratory results and liver biopsy showing evidence of hepatitis, and confirmed by detection of anti-HAV IgM antibodies. Because the mother had an acute icteric hepatitis A 1 week before delivery, and the viraemic phase of hepatitis A infection is very short, approximately 7 d, we suggest that the infant was infected by his mother, before birth.

[Hennekam syndrome]. / Syndrome de Hennekam.

UNLABELLED: Hennekam syndrome is a disorder comprising intestinal lymphangiectasia, facial anomalies and moderate mental retardation. Eight cases have been previously reported. CASE REPORT: A 17-month-old girl was admitted to hospital for peripheral edema. On physical examination, she presented with a normal mental development. Facial anomalies were noted including a flat face, depressed and broad nasal bridge, puffy eye lids, mild down-slanting palpebral fissures, hypertelorism, epicanthal folds, bulbous nasal tip, small mouth, and low set ears. A simian line and haemangiomas on the arms, trunk and left limb were also noted. There was no organomegaly. Laboratory investigations showed iron deficiency anemia, hypoproteinemia, hypogammaglobulinemia and an elevated level of alpha-1 antitrypsin excreted in the feces. Endoscopic investigation and the small bowel biopsy showed findings consistent with lymphangiectasia. The patient did well on 24 hour enteral nutrition including medium-chain triglyceride rich diet and infusion of human albumin. CONCLUSION: We have aimed to remind that Hennekam syndrome should be included in differential diagnosis when intestinal lymphangiectasia are associated with facial anomalies.

[Treatment of chronic viral hepatitis B in children with moderate doses of alpha interferon]. / Traitement de l'hépatite chronique à virus B de l'enfant par des doses modérées d'interféron alpha.

AIM: The alpha interferon treatment criteria have not been established in children with chronic hepatitis B. We report the results of a prospective study. METHODS: Between 1988-1992 14 children (2 girls and 12 boys) with chronic hepatitis B received 3 million U/m2 of interferon alpha three times a week for 6 months. All patients underwent a liver biopsy that showed a pattern of chronic active hepatitis. One patient had cirrhosis. Hepatitis B surface antigen, hepatitis Be antigen and hepatitis B virus DNA had been positive in the serum in all for at least 6 months and anti-delta antibodies were negative in all. Pretreatment aminotransferase levels were at least 1.5 times the upper limit of normal. RESULTS: After treatment patients were followed up for at least one year (mean: 21.5 +/- 8.3 months). At the end of treatment HBV DNA was negative in 13 out of 14 patients and reappeared in one; HBeAg seroconversion was observed in 11 patients with the appearance of anti-HBe antibodies. Six patients lost the HBs antigen within 1 to 14 months after treatment. Anti-HBs antibodies did not appear in any patients and aminotransferase level normalized in 13 patients. Thirteen patients underwent liver biopsy after treatment which showed improvement in 12. CONCLUSIONS: Treatment with alpha interferon at doses of 3 MU/m2 is effective in children with active hepatitis B. Long-term follow up is needed to evaluate the effectiveness of this therapy.