An innovative three-dimensional model of normal human skin to study the proinflammatory psoriatic effects of tumor necrosis factor-alpha and interleukin-17.

BACKGROUND: Among all cytokines involved in the pathogenesis and in the progression of psoriasis, Tumor Necrosis Factor (TNF)-alpha and interleukin (IL)-17 play a pivotal role. OBJECTIVE: The aim of the present study was to mimic a psoriatic microenvironment and to investigate the early effects of TNF-alpha and IL-17 in a three-dimensional model of organotypic normal human skin. METHODS: Human skin explants were obtained from plastic aesthetic surgery of healthy young women 20-40years old (n=7). The study was approved by the Institutional Review Board and written informed consent was obtained from all subjects. Bioptic fragments were cultured at the air-liquid interface overnight in a Transwell system and further divided before adding either 50ng/ml IL-17 or 100ng/ml TNF-alpha or a combination of both cytokines. For each subject, a control sample was cultured without any cytokine. Samples were harvested 24 or 48h after cytokine incubation. At both time points and for all cytokine treatments a bioptic fragment obtained from each patient was processed. Epidermal proliferation, expressions of terminal differentiation (keratin 10, K10, and 14, K14) and of intercellular adhesion (occludin for tight junctions and E-cadherin for adherens junctions) biomarkers were investigated by indirect immunofluorescence. RESULTS: IL-17 and TNF-alpha induced an early and statistically significant inhibition of keratinocyte proliferation (more than 80% compared with their respective controls). At 24h, the combination of both cytokines did not further reduce cell proliferation. Starting from 24h of incubation, a non-continuous occludin expression in the granular layer was observed after both IL-17 and TNF-alpha exposure. Immunolabelling for E-cadherin in adherens junctions, for K10 in the suprabasal layers, and for K14 in the basal layer was similar in all experimental groups and unaffected after cytokine treatment. CONCLUSIONS: These results suggest that in this experimental model IL-17 and TNF-alpha induced an early alteration of the homeostasis of the inner proliferative layer and of the upper granular layer, as shown by cell proliferation inhibition and occludin expression.

Influence on chondrogenesis of human osteoarthritic chondrocytes in co-culture with donor-matched mesenchymal stem cells from infrapatellar fat pad and subcutaneous adipose tissue.

Co-culture of mesenchymal stem cells (MSCs) and articular chondrocytes (ACs) has been proposed for autologous cartilage cell-based therapies, to overcome the issues associated to limited availability of articular chondrocytes (ACs). To evaluate the potentiality of a co-culture approach in aged osteoarthritic patients, MSCs from infrapatellar fat pad (IFP-MSCs) and knee subcutaneous adipose tissue (ASCs) were co-cultured with donor-matched osteoarthritic, expanded and cryopreserved, ACs in a 75%/25% ratio. Co-cultures were prepared also from nasal chondrocytes (NCs) to evaluate their possible use as an alternative to ACs. Pellets were differentiated for 14 days, using mono-cultures of each cell type as reference. Chondrogenic genes SOX9, COL2A1, ACAN were less expressed in co-cultures compared to ACs and NCs. Total GAGs content in co-cultures did not differ significantly from values predicted as the sum of each cell type contribution corrected for the co-culture ratio, as confirmed by histology. No significant differences were observed for GAGs/DNA in mono-cultures, demonstrating a reduced chondrogenic potential of ACs and NCs. In conclusion, a small percentage of expanded and cryopreserved ACs and NCs did not lead to IFP-MSCs and ASCs chondro-induction. Our results suggest that chondrogenic potential and origin of chondrocytes may play a relevant role in the outcome of co-cultures, indicating a need for further investigations to demonstrate their clinical relevance in the treatment of aged osteoarthritic patients.

Inside-out and outside-in organotypic normal human skin culture: JAK-STAT pathway is activated after pro-inflammatory psoriatic cytokine exposure.

The Janus kinases-signal transducers and activators of transcription (JAK-STAT) signalling pathway are a pleiotropic cascade that involves ligands such as cytokines, hormones, and growth factors. Among cytokines, interleukin (IL)-17, IL-22, IL-23, and tumour necrosis factor (TNF)-alpha play a pivotal role in psoriasis. We aimed at investigating in an organotypic experimental model of normal human skin (n = 7 women between 20-40 years old, non-smokers) the early, direct, and specific effects of IL-17, IL-22, IL-23, TNF-alpha and a combination of the four cytokines (Mix) on the JAK-STAT/pathway. The expression of the psoriatic marker keratin (K) 17 was analyzed by immunofluorescence and molecular techniques after exposure to IL-23 or Mix. The Mix elicited a strong K17 up-regulation in keratinocytes at 72 h, reinforcing the hypothesis of a synergistic effect of different cytokines. High levels of JAK1 and STAT3 activation were detected, suggesting the involvement of JAK1/STAT3 pathway in the upregulation of K17. As the present study in an organotypic model of human skin reports a variable expression of JAK-STAT upon different cytokine stimuli and most of the JAK inhibitors for the psoriasis treatment have proven to have a clinical efficacy, these observations have a relevance to better understand the mechanisms of JAK-inhibitors in the skin.

Enhanced biological performance of human adipose-derived stem cells cultured on titanium-based biomaterials and silicon carbide sheets for orthopaedic applications.

It is well known that the surface properties of biomaterials may affect bone-healing processes by modulating both cell viability and osteogenic differentiation. In this study we evaluated proliferation and osteogenic differentiation of human adipose-derived stem cells (hASCs) cultured on three prototypes of titanium disks and on thin layers of silicon carbide (SiC-PECVD), a material characterized by a high hardness and wear resistance. Our data indicated that all the tested surfaces supported cell growth, in particular, hASCs seeded on both titanium treated by a double-step etching process (TIT) and titanium modified by two Anodic Spark Deposition processes (TAA) grew better respect to the ones cultured on titanium obtained by KOH alkali etching process on TAA (TAAK). Furthermore, hASCs well colonized SiC-PECVD surface, showing a quite similar viability to cells cultured on plastic (PA). TIT and TAA better supported osteogenic differentiation of hASCs compared to PA, as shown by a marked increase of both alkaline phosphatase activity and calcified extracellular matrix deposition; in contrast TAAK did not positively affect hASCs differentiation. SiC-PECVD did not alter osteogenic differentiation of hASC cells: indeed, ALP and calcium deposition levels were comparable to those of cells cultured on plastic. Furthermore, we observed similar results testing hASCs either pre-differentiated for 14 days in osteogenic medium or directly differentiated on biomaterials. Our study suggests that modifications of titanium surface may improve osteo-integration of implant devices and that SiC-PECVD may represent a valid alternative for the coating of prosthetic devices to reduce wear and metallosis events.

Congenital ptosis and blefarophimosis: retrospective analysis of the effectiveness of correction with levator resection and frontalis suspension.

BACKGROUND: treatment of congenital ptoses and blepharophimoses relies on levator resections and frontalis suspensions. Several techniques of levator resection have been described in literature, some of them include tarsal resections and resections of the Müller muscle. Nevertheless a gold treatment have not been detected yet. Frontalis suspension is performed when levator muscle is not functional or when ptosis is severe. The suspension could be carried out with several materials: ePTFE, silicon rods, poly-propylene, nylon, braided poliester, but the preferable material is considered the autologous fascia lata. AIMS: this study was designate to determine retrospectively if the indications of surgery are correct, considering age, severity of pathology, relapses and complications. An analysis of demographic data and outcomes for each technique is performed. METHODS: in this study we analyze case series of 33 pediatric patients affected by congenital ptosis and blepharophimosis congenital syndrome, surgically treated from 2000 to 2008 in the ophtalmic pediatric surgery department at the Niguarda Hospital of Milan. A literature review was also performed. RESULTS: the mean age at presentation was 4.13. The diagnosis was precocious in most cases and often helped by some recognizable clinical signs: compensatory head posture (48.5%), anisometropia (36.4%), astigmatism (48.5%), strabismus (36.4%) and amblyopia (15.2%). Most of patients was treated with frontalis suspension (57.6%) and their age was significatively lower than patients treated with levator resection. No difference about complications and recurrence was reported between the two techniques. Complications and recurrence amount to 39.4%. CONCLUSION: these results are in line with other studies in literature. A precociuos treatment is able to reduce the incidence of amblyopia from 34% to 8%. The choice of the treatment (resection Vs suspension) has to consider the age of the patient, the severity of ptosis and avaibility of fascia lata. Nevertheless no significative difference in outcomes have been demonstrated between the two techniques.

Ten-year experience with craniofacial implants: clinical and experimental results.

The choice of bone substitute to be used in the cranio-maxillofacial districts raises several problems relating to the biocompatibility and mechanical resistance of the material. Over the past 10 years a total of 245 cranio-maxillofacial skeletal implants have been performed by the Department of Plastic Surgery at Niguarda Ca' Granda Hospital in Milan. The choice of material varied depending on the market availability of products and the experience acquired of both positive results and the onset of short-term and long-term complications. All patients were monitored with a minimum follow-up of 2 years and a maximum of 10 (mean 36 months). The following parameters were taken into account: complications linked to graft contamination, graft extrusion and decubitus, resorption times, degradable materials, resorption of underlying bone tissue and reactivity of the surrounding tissues. Studies on the cell cytohistotoxicity of materials used were performed in parallel by the cell culture laboratory at the Department of Plastic Surgery of Niguarda Hospital, using human keratinocyte and fibroblast cultures.

[Gynecomastia: diagnostic and surgical approach in the treatment of 61 patients]. / La ginecomastia: approccio diagnostico e chirurgico nel trattamento di 61 pazienti.

Gynaecomastia is a disease with a high incidence (approximately 65% of adult males between 15 and 40 Ys.). In this paper the authors present their experience about the medical and surgical treatment comparing different surgical techniques: adenomammectomy, liposuction and liposuction associated with adenomammectomy. 61 patients ageing 17-42 Ys. (54 with bilateral gynaecomastia and 7 with monolateral disease) were, treated in the Dept. of Plastic Surgery of the Niguarda Ca' Granda Hospital in Milan from 1985 up to 1995. 26 patients were treated with adenomammectomy; Suction assisted lipectomy was used alone in 4 cases and associated with adenomammectomy in 34 cases. The authors suggest that the associated method is the most effective, the aesthetic results being excellent with an important reduction of post-operative complications (mostly referred as haematoma, seroma).

Fire disasters in the twentieth century.

In the field of natural and man-made disasters, fire has played a predominant role. A report is presented of fire disasters in the twentieth century, with a chronological analysis of different worldwide typologies.

The use of propofol and remifentanil for the anaesthetic management of a super-obese patient.

Morbid obesity is defined as body mass index (BMI) > 35 kg.m(-2), and super-obesity as BMI > 55 kg.m(-2). We report the case of a 290-kg super-obese patient scheduled for open bariatric surgery. A propofol-remifentanil TCI (target controlled infusion) was chosen as the anaesthetic technique both for sedation during awake fibreoptic nasotracheal intubation and for maintenance of anaesthesia during surgery. Servin's weight correction formula was used for propofol. Arterial blood samples were taken at fixed time points to assess the predictive performance of the TCI system. A significant difference between measured and predicted plasma propofol concentrations was found. After performing a computer simulation, we found that predictive performance would have improved significantly if we had used an unadjusted pharmacokinetic set. However, in conclusion (despite the differences between measured and predicted plasma propofol concentrations), the use of a propofol-remifentanil TCI technique both for sedation during awake fibreoptic intubation and for Bispectral Index-guided propofol-remifentanil anaesthesia resulted in a rapid and effective induction, and operative stability and a rapid emergence, allowing rapid extubation in the operating room and an uneventful recovery.

[A case of Merkel carcinoma with facial localization]. / Un caso di carcinoma di Merkel a localizzazione facciale.

A case of Merkel carcinoma is reported. It concerns a 40-year-old lady, who had first noticed the lesion one year before she came to our observation. The authors point out the rarity of this tumour, whose not distinctive gross appearance (firm, sessile tumour with well circumscribed margins; hyperhemic overlaying skin; slowly increasing volume) may suggest other macroscopic diagnoses such as intradermal nevus or nodular basalioma. The authors also point out the usefulness of a prompt correct clinical diagnosis of this lesion, whose clinical behaviour may be highly aggressive.

The effects of scalp expansion on the cranial bone: a clinical, histological, and instrumental study.

The effects of chronic compression of the cranial bone due to progressive expansion of the scalp have been investigated. Ten patients were studied, 6 adults and 4 children, who were treated for congenital (microtia) or acquired (burns or traumatic) deformities by chronic expansion over a 2-month period. All underwent computed tomography scans of the expansion site prior to introduction of the expansion device, immediately before removal, and at 9 months after the operation. A case of postburn alopecia was lost to the study, because the patient, who had ultimated scalp expansion, did not return for flap advancement. Instead, she came back 3 months later, without the expander, which had been removed at another institution following an automobile accident. In this case, because of slower healing, we performed computed tomography scans 18 months postoperation. During the second procedure (expander removal and flap transposition), bone samples for histological examination were collected directly underneath and along the perimeter of the expanders. Macroscopically, the bone appeared thinned and had a reduced convexity. This reaction, although temporary, appeared more intense in the children and in the posttraumatic cases. Histological examination showed osteoclastic activation, bony hypotrophy, and reaction (deposition of osteoid matrix) under the device, with consequent bone resorption and remodeling. A marked hyperplasia with a hyperostotic reaction was observed around the expanders. At 9 months postoperation, in most cases, a complete normalization was confirmed by computed tomography scans. Expansion of target (fontanellar and sagittal) areas of the skull in children, as well as previous trauma to both scalp and skull should be taken into consideration as a risk factor. Further investigations are suggested.