Hyaluronic Acid Fillers, Needle Contamination by Fastidious Microorganisms, and Risk of Complications.

BACKGROUND: Complications are becoming ever more common with the increased use of hyaluronic acid (HA) fillers in aesthetic medicine. Complications due to needle contamination with fastidious microorganisms are no exception. OBJECTIVE: To perform, in a top Italian aesthetic medicine facility, what the authors think is the first monitoring program of microbial needle contamination of cross-linked HA gel fillers after the prefilled syringes with gel residues were stored for retouches after the first aesthetic procedure. METHODS: Needle contamination monitoring study, performed between January and November 2019, on 35 needles (caliber, 30 and 27 G) stored at 4°C in their resealed filler packages for possible retouch after a first aesthetic treatment involving the middle and lower facial thirds. Women's age: 35 to 70 years old. RESULTS: The search for contaminating agents of the 3 monitored bacterial contaminants ( Staphylococcus aureus , Streptococcus pyogenes , and anaerobes) as well as yeasts and molds always tested negative. In the days and months after treatment, no patients in post-treatment controls showed evidence of infection in the treated areas. CONCLUSION: The observational retrospective study confirms that good storage conditions, including monitored refrigeration, avoid the risk of contamination of partially used HA gel fillers by fastidious microorganisms.

An infrastructure for precision medicine through analysis of big data.

BACKGROUND: Nowadays, the increasing availability of omics data, due to both the advancements in the acquisition of molecular biology results and in systems biology simulation technologies, provides the bases for precision medicine. Success in precision medicine depends on the access to healthcare and biomedical data. To this end, the digitization of all clinical exams and medical records is becoming a standard in hospitals. The digitization is essential to collect, share, and aggregate large volumes of heterogeneous data to support the discovery of hidden patterns with the aim to define predictive models for biomedical purposes. Patients' data sharing is a critical process. In fact, it raises ethical, social, legal, and technological issues that must be properly addressed. RESULTS: In this work, we present an infrastructure devised to deal with the integration of large volumes of heterogeneous biological data. The infrastructure was applied to the data collected between 2010-2016 in one of the major diagnostic analysis laboratories in Italy. Data from three different platforms were collected (i.e., laboratory exams, pathological anatomy exams, biopsy exams). The infrastructure has been designed to allow the extraction and aggregation of both unstructured and semi-structured data. Data are properly treated to ensure data security and privacy. Specialized algorithms have also been implemented to process the aggregated information with the aim to obtain a precise historical analysis of the clinical activities of one or more patients. Moreover, three Bayesian classifiers have been developed to analyze examinations reported as free text. Experimental results show that the classifiers exhibit a good accuracy when used to analyze sentences related to the sample location, diseases presence and status of the illnesses. CONCLUSIONS: The infrastructure allows the integration of multiple and heterogeneous sources of anonymized data from the different clinical platforms. Both unstructured and semi-structured data are processed to obtain a precise historical analysis of the clinical activities of one or more patients. Data aggregation allows to perform a series of statistical assessments required to answer complex questions that can be used in a variety of fields, such as predictive and precision medicine. In particular, studying the clinical history of patients that have developed similar pathologies can help to predict or individuate markers able to allow an early diagnosis of possible illnesses.

In-vitro culture system for mesenchymal progenitor cells derived from waste human ovarian follicular fluid.

To characterize different cell populations in the human ovary, morphological and functional characteristics of cell populations collected during routine IVF procedures were studied. Cells obtained from follicular fluid grew in vitro under minimal medium conditions, without growth factor, including leukaemia-inhibiting factor. Morphological analysis revealed a heterogeneous cell population, with cells displaying a fibroblast-like, epithelial-like and also neuron-like features. Morpho-functional characteristics of fibroblast-like cells were similar to mesenchymal stem cells, and, in particular, were positive for mesenchymal stemness markers, including CD90, CD44, CD105, CD73, but negative for epithelial proteins, such as cytokeratins, CD34 and CD45 antigens. Cell proliferation activity at different times and colony-forming unit capability were evaluated, and multipotency of a subset of granulosa cells was established by in-vitro differentiation studies (e.g. osteogenic, chondrogenic and adipogenic differentiation). This study suggests that cells provided by mesenchymal plasticity can be easily isolated by waste follicular fluid, avoiding scraping of human ovaries, and cultivated in minimal conditions. Successful growth of such progenitor cells on three-dimensional cryogel scaffold provides the basis for future developments in tissue engineering. This culture system may be regarded as an experimental model in which biological behaviour is not influenced by specific growth factors.

Hypes and Hopes of Stem Cell Therapies in Dentistry: a Review.

One of the most exciting advances in life science research is the development of 3D cell culture systems to obtain complex structures called organoids and spheroids. These 3D cultures closely mimic in vivo conditions, where cells can grow and interact with their surroundings. This allows us to better study the spatio-temporal dynamics of organogenesis and organ function. Furthermore, physiologically relevant organoids cultures can be used for basic research, medical research, and drug discovery. Although most of the research thus far focuses on the development of heart, liver, kidney, and brain organoids, to name a few, most recently, these structures were obtained using dental stem cells to study in vitro tooth regeneration. This review aims to present the most up-to-date research showing how dental stem cells can be grown on specific biomaterials to induce their differentiation in 3D. The possibility of combining engineering and biology principles to replicate and/or increase tissue function has been an emerging and exciting field in medicine. The use of this methodology in dentistry has already yielded many interesting results paving the way for the improvement of dental care and successful therapies.

Dysplastic histogenesis of cartilage growth plate by alteration of sulphation pathway: a transgenic model.

Mutations in the diastrophic dysplasia sulphate transporter (dtdst) gene causes different forms of chondrodysplasia in the human. The generation of a knock-in mouse strain with a mutation in dtdst gene provides the basis to study developmental dynamics in the epiphyseal growth plate and long bone growth after impairment of the sulphate pathway. Our microscopical and histochemical data demonstrate that dtdst gene impairment deeply affects tissue organization, matrix structure, and cell differentiation in the epiphyseal growth plate. In mutant animals, the height of the growth plate was significantly reduced, according to a concomitant decrease in cell density and proliferation. Although the pathway of chondrocyte differentiation seemed complete, alteration in cell morphology compared to normal counterparts was detected. In the extracellular matrix, it we observed a dramatic decrease in sulphated proteoglycans, alterations in the organization of type II and type X collagen fibers, and premature onset of mineralization. These data confirm the crucial role of sulphate pathway in proteoglycan biochemistry and suggest that a disarrangement of the extracellular matrix may be responsible for the development of dtdts cartilage dysplasia. Moreover, we corroborated the concept that proteoglycans not only are structural components of the cartilage architecture, but also play a dynamic role in the regulation of chondrocyte growth and differentiation.

Generation of human epidermal constructs on a collagen layer alone.

Because engineered tissues are designed for clinical applications in humans, a major problem is the contamination of cocultures and tissues by allogenic molecules used to grow stem cells in vitro. The protocols that are commonly applied to generate epidermal equivalents in vitro require the use of irradiated murine fibroblasts as a feeder layer for keratinocytes. In this study, we report a simple procedure for growing human keratinocytes, isolated from adult skin, to generate an epidermal construct on a collagen layer alone. In this model, no human or murine feeder layers were used to amplify cell growth, and isolated keratinocytes were seeded directly at high cell density on the collagen-coated flasks or coverslips in an epithelial growth medium containing low calcium concentration. Morphological, immunochemical, and cytokinetic features of epithelial colonies grown on the collagen layer were typical of keratinocytes and were comparable with those reported for keratinocytes grown on a feeder layer. The stratification of keratinocytes generated 3-dimensional synthetic constructs displaying a tissue architecture comparable with that of natural epidermis. Epithelial cells expressed specific markers of keratinocyte terminal differentiation, including involucrin and filaggrin. Nevertheless, the number of cell layers was lower than in natural skin, and electron microscopical analysis revealed that the overall organization of these layers was poor compared with natural epidermis, including the formation of junctional complexes, basement membrane, and keratinization. The lack of epithelial-mesenchymal interactions that occur during skin histogenesis may account for such an incomplete maturation of epidermal constructs.

Cell kinetic analysis in artificial skin using immunochemical methods.

Cell kinetic studies provide important information on histogenesis in vivo and in vitro. In this regard, specific antibodies to cell cycle-related antigens have been raised and characterized, thus permitting the study of cell kinetics using immunochemical methods. Recent advances in culture technology permitted the generation of human skin equivalents in vitro. We here provide detailed practical procedures for the study of epidermal cell kinetics in a model of artificial skin using immunohistochemistry and flow cytometry. The combined application of both techniques allows a precise detection of tissue growth sites and a quantitative assessment of cell growth. Moreover, simultaneous analysis of differentiation markers and proliferation antigens may be useful to understand molecular mechanisms that regulate tissue growth and development.

The suborbicularis oculi fat (SOOF) and the fascial planes: has everything already been explained?

IMPORTANCE: During anatomic and surgical dissections, a connection was seen between the superficial layer of the deep temporal fascia and the prezygomatic area. These findings were in contrast to previous evaluations. This study defines this connection, which is important to understand from both surgical and anatomic standpoints. OBJECTIVE: To define the connection between the superficial layer of the deep temporal fascia and the prezygomatic area and demonstrate the presence of a deep fascial layer in the midface. DESIGN AND SETTING: Anatomical study performed at the Laboratoire d'Anatomie de la Faculté de Médecine de Nice, Sophia Antipolis, France; at the Centre du Don des Corps de l'Université Paris Descartes, Paris, France; and at the Department of Experimental Medicine, Histology, and Embryology Unit of the University of Pavia, Pavia, Italy. Twenty-four hemifaces of 14 white cadavers were dissected to define the relationship between deep temporal fascia and the midface. Four biopsy samples were harvested for histologic analysis. MAIN OUTCOMES AND MEASURES: Dissection of 24 hemifaces from the fresh cadavers revealed the following findings. There is a connection of the deep fascia of the temple (superficial layer of deep temporal fascia) to the midface that divides the fat deep to the orbicularis muscle into 2 layers. One layer of fat is the so-called suborbicularis oculi fat (SOOF), which is superficial to the deep fascia, and the other layer of fat (preperiosteal) is deep to the deep fascia and adherent to malar bone. These findings are in contrast to previous anatomical findings. RESULTS In 12 hemifaces, the superficial layer of the deep temporal fascia directly reached the prezygomatic area as a continuous fascial layer. In 16 hemifaces, the superficial sheet of the deep temporal fascia inserted at the level of the zygomatic and lateral orbital rim and continued as a deep fascial layer over the prezygomatic area. In all specimens, a deep fascial layer was present in the prezygomatic-infraorbital area. This deep fascial layer is adherent to the muscles of the infraorbital area, and it divided the fat located deep to the orbicularis oculi muscle into 2 layers: the SOOF and a deeper layer. Histologic examination of the biopsy samples confirmed these findings. CONCLUSIONS AND RELEVANCE: This study demonstrates the existence of a deep fascial layer in the midface. This fascia is connected to the superficial layer of the deep temporal fascia, and it divides the fat deep to the orbicularis oculi muscle into 2 layers. This new finding carries interesting implications related to the classic concept of the superficial musculoaponeurotic system. LEVEL OF EVIDENCE: NA.

Long-term miR-669a therapy alleviates chronic dilated cardiomyopathy in dystrophic mice.

BACKGROUND: Dilated cardiomyopathy (DCM) is a leading cause of chronic morbidity and mortality in muscular dystrophy (MD) patients. Current pharmacological treatments are not yet able to counteract chronic myocardial wastage, thus novel therapies are being intensely explored. MicroRNAs have been implicated as fine regulators of cardiomyopathic progression. Previously, miR-669a downregulation has been linked to the severe DCM progression displayed by Sgcb-null dystrophic mice. However, the impact of long-term overexpression of miR-669a on muscle structure and functionality of the dystrophic heart is yet unknown. METHODS AND RESULTS: Here, we demonstrate that intraventricular delivery of adeno-associated viral (AAV) vectors induces long-term (18 months) miR-669a overexpression and improves survival of Sgcb-null mice. Treated hearts display significant decrease in hypertrophic remodeling, fibrosis, and cardiomyocyte apoptosis. Moreover, miR-669a treatment increases sarcomere organization, reduces ventricular atrial natriuretic peptide (ANP) levels, and ameliorates gene/miRNA profile of DCM markers. Furthermore, long-term miR-669a overexpression significantly reduces adverse remodeling and enhances systolic fractional shortening of the left ventricle in treated dystrophic mice, without significant detrimental consequences on skeletal muscle wastage. CONCLUSIONS: Our findings provide the first evidence of long-term beneficial impact of AAV-mediated miRNA therapy in a transgenic model of severe, chronic MD-associated DCM.

Growth and stratification of epithelial cells in minimal culture conditions.

Biological risk management is required in modern tissue engineering. Particular attention should be paid to the culture medium and the scaffold used. In this perspective, it is important to define minimal culture conditions which allow proper growth and differentiation of epithelial cells in vitro. We propose a simple experimental system which permits the generation of three-dimensional epidermal constructs using a collagen layer as a scaffold mimicking the entire dermal tissue and without the need of any feeder layer. Although showing significant differences compared to natural epidermis, these epidermal constructs appear useful to study keratinocyte differentiation and epidermis histogenesis.

Cell-cycle inhibition by Helicobacter pylori L-asparaginase.

Helicobacter pylori (H. pylori) is a major human pathogen causing chronic gastritis, peptic ulcer, gastric cancer, and mucosa-associated lymphoid tissue lymphoma. One of the mechanisms whereby it induces damage depends on its interference with proliferation of host tissues. We here describe the discovery of a novel bacterial factor able to inhibit the cell-cycle of exposed cells, both of gastric and non-gastric origin. An integrated approach was adopted to isolate and characterise the molecule from the bacterial culture filtrate produced in a protein-free medium: size-exclusion chromatography, non-reducing gel electrophoresis, mass spectrometry, mutant analysis, recombinant protein expression and enzymatic assays. L-asparaginase was identified as the factor responsible for cell-cycle inhibition of fibroblasts and gastric cell lines. Its effect on cell-cycle was confirmed by inhibitors, a knockout strain and the action of recombinant L-asparaginase on cell lines. Interference with cell-cycle in vitro depended on cell genotype and was related to the expression levels of the concurrent enzyme asparagine synthetase. Bacterial subcellular distribution of L-asparaginase was also analysed along with its immunogenicity. H. pylori L-asparaginase is a novel antigen that functions as a cell-cycle inhibitor of fibroblasts and gastric cell lines. We give evidence supporting a role in the pathogenesis of H. pylori-related diseases and discuss its potential diagnostic application.

Cost-effectiveness analysis for trigeminal neuralgia: Cyberknife vs microvascular decompression.

BACKGROUND/AIMS: We present the preliminary results of a cost-effectiveness analysis of cyberknife radiosurgery (CKR) versus microvascular decompression (MVD) for patients with medically unresponsive trigeminal neuralgia. METHODS: Direct healthcare costs from hospital's perspective attributable to CKR and MVD were collected. Pain level caused by trigeminal neuralgia was measured through the Barrow Neurological Institute pain intensity scoring criteria, at admission and after an average of 6 months follow-up. RESULTS: 20 patients for both arms were enrolled, for a total of 40 patients. The two procedures resulted equally effective at 6 month follow-up, with different resources consumption: CKR reducing hospital costs by an average of 34% per patient. The robustness of these results was confirmed in appropriate sensitivity analyses. CONCLUSION: CKR resulted to be a cost-saving alternative compared with the surgical intervention.

Expression of p63 transcription factor in ectoderm-derived oral tissues.

The p63 gene encodes six splice variants expressed with transactivating or dominant-negative activities. Animal studies with p63 -/- mutants have suggested that p63 is important for proper development of several organs, including tooth and salivary gland. Moreover, mutations of p63 have been detected in patients affected by ectrodactyly, ectodermal dysplasia and facial clefts. To clarify the role of p63 in craniofacial development, we have studied the localization of p63 protein in human and rat oral tissues using immunohistochemistry. p63 immunostaining was identified in the enamel organ, oral epithelium and developing salivary glands. All compartments of the enamel organ were immunolabelled, whereas only basal and some suprabasal cells of the oral epithelium were stained. Ectomesenchyme-derived cells, including pulp cells, odontoblasts, bone cells and chondrocytes, were negative. The staining pattern was identical in human and rat tissues. These data lend support to the hypothesis that p63 is involved in growth and differentation of ectoderm-derived oral tissues and may be useful to clarify molecular and developmental aspects observed in animal knockout experiments and human syndromes related to p63 gene alteration.