IL-17 and IL-22 are pivotal cytokines to delay wound healing of S. aureus and P. aeruginosa infected skin.

Introduction: Although the presence of pathogens in skin wounds is known to delay the wound healing process, the mechanisms underlying this delay remain poorly understood. In the present study, we have investigated the regulatory role of proinflammatory cytokines on the healing kinetics of infected wounds. Methods: We have developed a mouse model of cutaneous wound healing, with or without wound inoculation with Staphylococcus aureus and Pseudomonas aeruginosa, two major pathogens involved in cutaneous wound bacterial infections. Results: Aseptic excision in C57BL/6 mouse skin induced early expression of IL-1ß, TNFα and Oncostatin M (OSM), without detectable expression of IL-22 and IL-17A/F. S. aureus and P. aeruginosa wound inoculation not only increased the expression of IL-1ß and OSM, but also induced a strong cutaneous expression of IL-22, IL-17A and IL-17F, along with an increased number of infiltrating IL-17A and/or IL-22-producing γδ T cells. The same cytokine expression pattern was observed in infected human skin wounds. When compared to uninfected wounds, mouse skin infection delayed the wound healing process. Injection of IL-1α, TNFα, OSM, IL-22 and IL-17 together in the wound edges induced delayed wound healing similar to that induced by the bacterial infection. Wound healing experiments in infected Rag2KO mice (deficient in lymphocytes) showed a wound healing kinetic similar to uninfected Rag2KO mice or WT mice. Rag2KO infected-skin lesions expressed lower levels of IL-17 and IL-22 than WT, suggesting that the expression of these cytokines is mainly dependent on γδ T cells in this model. Wound healing was not delayed in infected IL-17R/IL-22KO, comparable to uninfected control mice. Injection of recombinant IL-22 and IL-17 in infected wound edges of Rag2KO mice re-establish the delayed kinetic of wound healing, as in infected WT mice. Conclusion: These results demonstrate the synergistic and specific effects of IL-22 and IL-17 induced by bacterial infection delay the wound healing process, regardless of the presence of bacteria per se. Therefore, these cytokines play an unexpected role in delayed skin wound healing.

Observance of preoperative clear fluid fasting in pediatric anesthesia: oral and written information versus text message information. A before-and-after study.

INTRODUCTION: Prolonged fasting before surgery is common in pediatrics. In the literature, it is responsible for hypotension, irritability and postoperative nausea and vomiting. Despite clear instructions given during the preanesthetic consultation, fasting rules are respected in only 30%-40% of cases. We aimed to evaluate the benefit of sending a text message the day before surgery to improve the parents' observance of fasting rules. METHODS: We conducted a before-and-after study at the University Hospital of Poitiers. From August to October 2018, 172 parents of children under 15 years of age scheduled for all types of surgery were enrolled into two groups according to the period: the control group with parents receiving information on preoperative fasting rules during the preanesthetic consultation several days before surgery, and the text message group, receiving the same information during consultation plus a text message the day before the surgery. RESULTS: There was a difference in observance of clear fluid fasting instructions (between 2 and 3 h before the admission at hospital) in favor of the text message group 33% versus 92% OR 29.2 (10.9-95.2) p < 0.001, and in average fasting time for clear fluids 8.7 h ± 4.8 h vs. 4.3 h ± 2.4 h (p < 0.001). CONCLUSION: Sending of a reminder text message to the parents the day before the surgery resulted in a significant increase in observance of fasting rules in children undergoing scheduled surgery.

Development of a new model of reconstituted mouse epidermis and characterization of its response to proinflammatory cytokines.

The development of three-dimensional models of reconstituted mouse epidermis (RME) has been hampered by the difficulty to maintain murine primary keratinocyte cultures and to achieve a complete epidermal stratification. In this study, a new protocol is proposed for the rapid and convenient generation of RME, which reproduces accurately the architecture of a normal mouse epidermis. During RME morphogenesis, the expression of differentiation markers such as keratins, loricrin, filaggrin, E-cadherin and connexins was followed, showing that RME structure at day 5 was similar to those of a normal mouse epidermis, with the acquisition of the natural barrier function. It was also demonstrated that RME responded to skin-relevant proinflammatory cytokines by increasing the expression of antimicrobial peptides and chemokines, and inhibiting epidermal differentiation markers, as in the human system. This new model of RME is therefore suitable to investigate mouse epidermis physiology further and opens new perspectives to generate reconstituted epidermis from transgenic mice.

Interleukin-17A-induced production of acute serum amyloid A by keratinocytes contributes to psoriasis pathogenesis.

BACKGROUND: Acute-serum Amyloid A (A-SAA), one of the major acute-phase proteins, is mainly produced in the liver but extra-hepatic synthesis involving the skin has been reported. Its expression is regulated by the transcription factors NF-κB, C/EBPß, STAT3 activated by proinflammatory cytokines. OBJECTIVES: We investigated A-SAA synthesis by resting and cytokine-activated Normal Human Epidermal Keratinocytes (NHEK), and their inflammatory response to A-SAA stimulation. A-SAA expression was also studied in mouse skin and liver in a model mimicking psoriasis and in the skin and sera of psoriatic and atopic dermatitis (AD) patients. METHODS: NHEK were stimulated by A-SAA or the cytokines IL-1α, IL-17A, IL-22, OSM, TNF-α alone or in combination, previously reported to reproduce features of psoriasis. Murine skins were treated by imiquimod cream. Human skins and sera were obtained from patients with psoriasis and AD. A-SAA mRNA was quantified by RT qPCR. A-SAA proteins were dosed by ELISA or immunonephelemetry assay. RESULTS: IL-1α, TNF-α and mainly IL-17A induced A-SAA expression by NHEK. A-SAA induced its own production and the synthesis of hBD2 and CCL20, both ligands for CCR6, a chemokine receptor involved in the trafficking of Th17 lymphocytes. A-SAA expression was increased in skins and livers from imiquimod-treated mice and in patient skins with psoriasis, but not significantly in those with AD. Correlations between A-SAA and psoriasis severity and duration were observed. CONCLUSION: Keratinocytes could contribute to psoriasis pathogenesis via A-SAA production, maintaining a cutaneous inflammatory environment, activating innate immunity and Th17 lymphocyte recruitment.

Oncostatin M overexpression induces skin inflammation but is not required in the mouse model of imiquimod-induced psoriasis-like inflammation.

Oncostatin M (OSM) has been reported to be overexpressed in psoriasis skin lesions and to exert proinflammatory effects in vitro on human keratinocytes. Here, we report the proinflammatory role of OSM in vivo in a mouse model of skin inflammation induced by intradermal injection of murine OSM-encoding adenovirus (AdOSM) and compare with that induced by IL-6 injection. Here, we show that OSM potently regulates the expression of genes involved in skin inflammation and epidermal differentiation in murine primary keratinocytes. In vivo, intradermal injection of AdOSM in mouse ears provoked robust skin inflammation with epidermal thickening and keratinocyte proliferation, while minimal effect was observed after AdIL-6 injection. OSM overexpression in the skin increased the expression of the S100A8/9 antimicrobial peptides, CXCL3, CCL2, CCL5, CCL20, and Th1/Th2 cytokines, in correlation with neutrophil and macrophage infiltration. In contrast, OSM downregulated the expression of epidermal differentiation genes, such as cytokeratin-10 or filaggrin. Collectively, these results support the proinflammatory role of OSM when it is overexpressed in the skin. However, OSM expression was not required in the murine model of psoriasis induced by topical application of imiquimod, as demonstrated by the inflammatory phenotype of OSM-deficient mice or wild-type mice treated with anti-OSM antibodies.

Foreskin-isolated keratinocytes provide successful extemporaneous autologous paediatric skin grafts.

Severe burns in children are conventionally treated with split-thickness skin autografts or epidermal sheets. However, neither early complete healing nor quality of epithelialization is satisfactory. An alternative approach is to graft isolated keratinocytes. We evaluated paediatric foreskin and auricular skin as donor sources, autologous keratinocyte transplantation, and compared the graft efficiency to the in vitro capacities of isolated keratinocytes to divide and reconstitute epidermal tissue. Keratinocytes were isolated from surgical samples by enzymatic digestion. Living cell recovery, in vitro proliferation and epidermal reconstruction capacities were evaluated. Differentiation status was analysed, using qRT-PCR and immunolabelling. Eleven children were grafted with foreskin-derived (boys) or auricular (girls) keratinocyte suspensions dripped onto deep severe burns. The aesthetic and functional quality of epithelialization was monitored in a standardized way. Foreskin keratinocyte graft in male children provides for the re-epithelialization of partial deep severe burns and accelerates wound healing, thus allowing successful wound closure, and improves the quality of scars. In accordance, in vitro studies have revealed a high yield of living keratinocyte recovery from foreskin and their potential in terms of regeneration and differentiation. We report a successful method for grafting paediatric males presenting large severe burns through direct spreading of autologous foreskin keratinocytes. This alternative method is easy to implement, improves the quality of skin and minimizes associated donor site morbidity. In vitro studies have highlighted the potential of foreskin tissue for graft applications and could help in tissue selection with the prospect of grafting burns for girls.

Study of proliferation and 3D epidermal reconstruction from foreskin, auricular and trunk keratinocytes in children.

OBJECTIVE: Severe burns in children are conventionally treated with split-thickness skin autografts or epidermal sheets. An alternative approach is to graft isolated keratinocytes. We evaluated foreskin and other anatomic sites as donor sources for autologous keratinocyte graft in children. We studied in vitro capacities of isolated keratinocytes to divide and reconstitute epidermal tissue. METHODS: Keratinocytes were isolated from foreskin, auricular skin, chest and abdominal skin by enzymatic digestion. Living cell recovery, in vitro proliferation, epidermal reconstruction capacities and differentiation status were analyzed. RESULTS: In vitro studies revealed the higher yield of living keratinocyte recovery from foreskin and higher potential in terms of proliferative capacity, regeneration and differentiation. Cultured keratinocytes from foreskin express lower amounts of differentiation markers than those isolated from trunk and ear. Histological analysis of reconstituted human epidermis derived from foreskin and inguinal keratinocytes showed a structured multilayered epithelium, whereas those obtained from ear pinna-derived keratinocytes were unstructured. CONCLUSION: Our studies highlight the potential of foreskin tissue for autograft applications in boys. A suitable alternative donor site for autologous cell transplantation in female paediatric burn patients remains an open question in our department. We tested the hypothesis that in vitro studies and RHE reconstructive capacities of cells from different body sites can be helpful to select an optimal site for keratinocyte isolation before considering graft protocols for girls.

Epidermal healing in burns: autologous keratinocyte transplantation as a standard procedure: update and perspective.

BACKGROUND: Treatment of burned patients is a tricky clinical problem not only because of the extent of the physiologic abnormalities but also because of the limited area of normal skin available. METHODS: Literature indexed in the National Center (PubMed) has been reviewed using combinations of key words (burns, children, skin graft, tissue engineering, and keratinocyte grafts). Articles investigating the association between burns and graft therapeutic modalities have been considered. Further literature has been obtained by analysis of references listed in reviewed articles. RESULTS: Severe burns are conventionally treated with split-thickness skin autografts. However, there are usually not enough skin donor sites. For years, the question of how covering the wound surface became one of the major challenges in clinical research area and several procedures were proposed. The microskin graft is one of the oldest methods to cover extensive burns. This technique of skin expansion is efficient, but results remain inconsistent. An alternative is to graft cultured human epidermal keratinocytes. However, because of several complications and labor-intensive process of preparing grafts, the initial optimism for cultured epithelial autograft has gradually declined. In an effort to solve these drawbacks, isolated epithelial cells from selecting donor site were introduced in skin transplantation. CONCLUSIONS: Cell suspensions transplanted directly to the wound is an attractive process, removing the need for attachment to a membrane before transfer and avoiding one potential source of inefficiency. Choosing an optimal donor site containing cells with high proliferative capacity is essential for graft success in burns.

Mutations in the human UBR1 gene and the associated phenotypic spectrum.

Johanson-Blizzard syndrome (JBS) is a rare, autosomal recessive disorder characterized by exocrine pancreatic insufficiency, typical facial features, dental anomalies, hypothyroidism, sensorineural hearing loss, scalp defects, urogenital and anorectal anomalies, short stature, and cognitive impairment of variable degree. This syndrome is caused by a defect of the E3 ubiquitin ligase UBR1, which is part of the proteolytic N-end rule pathway. Herein, we review previously reported (n = 29) and a total of 31 novel UBR1 mutations in relation to the associated phenotype in patients from 50 unrelated families. Mutation types include nonsense, frameshift, splice site, missense, and small in-frame deletions consistent with the hypothesis that loss of UBR1 protein function is the molecular basis of JBS. There is an association of missense mutations and small in-frame deletions with milder physical abnormalities and a normal intellectual capacity, thus suggesting that at least some of these may represent hypomorphic UBR1 alleles. The review of clinical data of a large number of molecularly confirmed JBS cases allows us to define minimal clinical criteria for the diagnosis of JBS. For all previously reported and novel UBR1 mutations together with their clinical data, a mutation database has been established at LOVD.

Infantile hypertrophic pyloric stenosis: are viruses involved?

Infantile hypertrophic pyloric stenosis (IHPS) is characterized by abnormal thickening of the internal circular muscle layer. IHPS is known to be due to a combination of genetic and environmental factors, but its precise causes and pathophysiology are poorly understood. The objective of the study is to determine the prevalence of the principal viruses targeting the respiratory and digestive tracts in children with IHPS. Nasopharyngeal fluids, stools, vomit, and surgical pyloric muscle fragments and swabs were tested by cell culture, viral antigen assay and PCR. IHPS was diagnosed in 23 boys and 8 girls with a mean (± SD) age of 42 ± 15 days (range 20-88 days). There was no seasonal pattern of diagnosis. Twenty-two children (71%) lost weight (mean 246 ± 164 g, range 30-600 g) after the onset of vomiting, and five (16.1%) were dehydrated. Seven (22.6%) infants had been exposed to an infectious contact within 15 days before admission, and one on the day of admission (3.2%). Ear, nose and throat samples and pyloric muscle specimens were negative for all the viruses tested. An adenovirus type 3 was recovered from one stool sample, and RT-PCR was positive for an enterovirus on one vomit sample. This study suggests that the principal viruses targeting the respiratory and digestive tracts are not responsible for IHPS.

Wandering spleen in children: multicenter retrospective study.

Wandering spleen in children is a rare condition. The diagnosis is difficult, and any delay can cause splenic ischemia. An epidemiologic, semiological, and surgical diagnosis questionnaire on incidence of wandering spleen in children was sent to several French surgical teams. We report the results of this multicenter retrospective study. Fourteen cases (6 girls, 8 boys) were reported between 1984 and 2009; the age range varies between 1-day-old and 15 years; 86% were seen in the emergency department. Ninety-three percent had diffuse abdominal pain. For 57% of the cases, it was their first symptomatic episode of this type. No diagnosis was established based on the clinical results alone. All patients had presurgical imaging diagnosis. Open surgery was performed on 64% cases. Forty-three had splenectomy for splenic ischemia. Thirty-six percent had splenopexy, 14% had laparoscopic gastropexy, and 7% had spleen repositioning and regeneration. Complications were noted in 60% of the cases resulting in postsplenopexy splenic ischemia. Early diagnosis and surgery are the best guarantee for spleen preservation. Even if the choice of one technique, splenopexy or gastropexy, can be argued, gastropexy has the advantage of avoiding splenic manipulation and restoring proper physiologic anatomy. When there is no history of abdominal surgery, laparoscopy surgery seems the best procedure.

Growth in infants in the first two years of life after neonatal repair for unilateral cleft lip and palate.

OBJECTIVE: To evaluate the growth during the first two years of life in infants after unilateral cleft lip and palate neonatal repair. METHOD: All mature infants with nonsyndromic unilateral cleft lip and palate (NSUCLP) born between 2004 and 2007 were included. Information concerning growth was collected. Weight and length at birth, 6, 12, 18 and 24 months of age measurements and data regarding feeding were obtained. RESULTS: Weight and length at birth, 6, 12, 18 and 24 months of age were identical with reference curve values. Children with NSUCLP showed a normal growth at two years. The weight curves lie between 5th and the 50th percentile for girls and between 10th and higher than the 97th percentile for boys. The height curves lie between -1 Standard Deviation and +1 Standard Deviation for girls and 0 and +2 Standard Deviation for boys. CONCLUSION: Feeding difficulties are reported in infants with cleft lip and/or palate CLP/CP. However, the growth in children with NSUCLP and after neonatal cleft lip repair is identical with reference curve values.

[Neonatal computed tomographic maxillary explorations in children with unilateral cleft lip and palate]. / Etudes tomodensitométriques néonatales de l'os maxillaire dans les fentes labiopalatines unilatérales.

The aim was to examine the bony maxillary structures by computed tomographic measurements in newborns with unilateral cleft lip and palate before cheiloplasty. Analysis of maxillary bone was performed and size parameters were measured by computed-tomographic analysis in 12 infants with unilateral cleft lip and palate. We compared the bony maxillary length and the bony maxillary width between the cleft side and the healthy side. For eight patients, the bony maxillary length was different between the cleft side and the healthy side. For three patients, the bony maxillary width was different between the incisor alveolar structure in the cleft side and the healthy side. For six patients, the bony maxillary width was different between the canine alveolar structure in the cleft side and the healthy side. We noted an asymmetry without hypoplasy in bony maxillary structure in newborns before cheiloplasty. The data can serve as the starting point for a control and later evaluation on the efficiency of different therapeutic approaches of alveolar and maxillary development in children with cleft lip and palate.

Solitary intestinal fibromatosis associated with congenital ileal atresia.

We report for the first time an association between congenital solitary intestinal fibromatosis and intestinal atresia. The spindle cell proliferation showed a high apoptotic index contrasting with a low proliferation rate, suggesting that the tumor may have undergone focal and spontaneous regression, leading to intestinal atresia.

Early repair for infants with cleft lip and nose.

OBJECTIVE: To study the results 10 years after early surgical cleft lip and nose repair. PATIENTS AND METHODS: We present the outcome of 123 early cleft lip repairs whose condition was managed in a multidisciplinary team according to a strict protocol. We give the observation results of operations of a single surgeon's neonatal surgery over a 10-year period in term of aesthetic assessments and dental arch relationships. One hundred and twenty-three patients were operated on during the first 4 weeks of life; a subgroup of 40 child had been operated on at a week or less of age. RESULTS: The results show good aesthetic assessments and dental relationships, with facial growth appropriate for the age. CONCLUSIONS: We are currently encouraging early cleft lip and nose repair in the full-term baby as the good method of management of newborns with cleft.

Surgical treatment of haemangioma in infants.

Haemangiomas usually can be identified by their clinical course. They are characterised by presentation at birth or shortly thereafter, and a rapid proliferative phase over the first 12 months. The haemangioma then usually stabilises and slowly involutes over a period of 5-7 years. For a long time, surgery has been limited to complicated cases, and correcting after-effects following involution. Nevertheless, aesthetic, psychological or functional prejudices may justify early surgery. We conducted a retrospective study of patients treated between 1995 and 2001. A total of 31 patients with facial and cervical haemangiomas were studied. For each, the type of lesion and its topography, age and operative indications, surgery, postoperative complications and aesthetic and functional results have been considered. Thirty-one haemangiomas were operated. The average age was 30 months (1-60 months). After an average follow-up of 3 years, the results were very good in 20%, good in 66%, and fair in 14% of cases. Early curative surgery of haemangioma before spontaneous involution, and before school-age is justified because of social and psychological considerations in infants and their family.

[Vesicoureteral reflux: diagnosis and management in children]. / Reflux vésico-urétéral: diagnostic et prise en charge chez l'enfant.

OBJECTIVE: Based on a retrospective study of children followed for vesicoureteric reflux, the authors evaluated the role of antenatal diagnosis in the detection and global management of the most frequent uropathy observed in children. MATERIALS AND METHODS: The case files of 180 children followed for reflux over a 4-year period were reviewed. The diagnosis of reflux was based on retrograde cystography. The authors tried to define the main indications for the renal and urinary tract assessment and defined the grade and type of reflux, as well as the therapeutic indications and results. 180 children (105 girls and 75 boys) with vesicoureteric reflux were observed during the study period. The mean age at the time of diagnosis was 26.75 months. RESULTS: Vesicoureteric reflux was usually diagnosed in a context of acute pyelonephritis in 139 children (77%);, at the time of the first episode in 84% of cases. Antenatal diagnosis of reflux during screening for dilated renal pelvis was the second most frequent modality (29 children, 16% of cases). The reflux involved a single urinary tract in 163 children and a duplicated tract in 17 children. Reflux was bilateral in 43% of cases. Reflux was less than grade II in 66% of cases, but an inverse proportion was observed in the case of antenatal diagnosis (62%). 55% of cases were treated surgically and 45% were treated medically. Follow-up of the children showed a low recurrent pyelonephritis rate that was similar in the two groups. CONCLUSION: Acute pyelonephritis remains the leading mode of discovery of vesicoureteric reflux and renal ultrasonography combined with retrograde cystography after a first episode of renal infection is therefore recommended. Antenatal screening may also reveal vesicoureteric reflux. It is difficult to summarize the therapeutic indications for reflux, as the treatment of vesicoureteric reflux is related to the child's age and sex, the grade of reflux, the clinical repercussions of the reflux and certain environmental factors, such as the family and social status.

[Laparoscopic treatment of the nonpalpable testis. Results]. / Traitement laparoscopique des testicules impalpables. Résultats.

OBJECTIVE: The purpose of laparoscopy in the management of the non palpable testis is to provide information regarding testicular presence and location to facilitate overall surgical management. MATERIALS AND METHODS: We report our experience with laparoscopic orchiopexy to treat 39 nonpalpable testes in 32 patients, patient age ranged from 2.3 years to 14 years (average 4.18 years). We retrospectively reviewed the medical records of all patients for a 5-year period. RESULTS: At laparoscopy 11 testes were at the internal inguinal ring, these patients underwent one-stage laparoscopic orchiopexy without division of the spermatic vessels. 18 high intra-abdominal testes underwent two-stage Fowler-Stephens orchiopexy. One Patient underwent laparoscopically assisted orchiectomy for atrophy, 9 testes were absent. At follow-up 6, 12 and 24 months after 26 of 29 (89%) testes are without atrophy, and in acceptable scrotal position. CONCLUSIONS: The low incidence of complications and 89% success rate underscore the feasibility of laparoscopic orchiopexy. It is our procedure of choice for the management and treatment of nonpalpable testis.