Laser induced autofluorescence studies of animal skin used in modeling of human cutaneous tissue spectroscopic measurements

BACKGROUND/PURPOSE: Laser-induced autofluorescence spectroscopy provides excellent possibilities for medical diagnostics of different tissue pathologies including cancer. However, to create the whole picture of pathological changes, investigators collect spectral information from patients in vivo or they study different tumor models to obtain objective information for fluorescent properties of every kind of healthy and diseased tissue. Therefore, it is very important to find the most appropriate, and close to the human skin, animal samples from the fluorescence point of view, which will allow the extrapolation of the animal data to human spectroscopic diagnostics. METHODS: In the present work, we examined the autofluorescence properties of different animal skin tissues, which are considered as the most common skin models. A nitrogen laser was used as an excitation source. Samples of healthy mouse, chicken and pig skin in vivo and/or ex vivo were studied and were compared with results obtained from investigations of healthy human skin in vivo. RESULTS AND CONCLUSION: Specific features of the recorded spectra are discussed and the possible origin of the obtained fluorescence signals is proposed. Quantitative evaluation of data extrapolation for each skin type is also depicted.     

The Histological Basis of Frank’s Sign

Frank’s sign is a diagonal crease of the ear lobe, supposedly related to cardiac pathology, and has strongly been associated with coronary artery atherosclerosis. A total of 45 consecutive adult patients referred for autopsy in a one-and-a-half-year period were extensively studied. Samples from both the ear lobes were obtained for histopathology, as well as cardiac samples from all four cardiac compartments. When compared patients with Frank’s sign and those without it had no statistical difference in age (p = 0.0575). There was however a statistically significant increased cardiac weight (p = 0.0005), left ventricular wall thickness (p = 0.0002), and right ventricular wall thickness (p = 0.0043). Histopathology obtained from the ear lobes revealed myoelastofibrosis in an arterial vessel, located at the base of the crease, diffuse fibrosis, and Wallerian-like degeneration, with eosinophilic inclusions in the peripheral nerves. These changes suggest a time-related progression of the crease-associated changes. Our data suggest a significant correlation between the morphological changes of the myocardium and the presence of the ear lobe creases, with arterial myoelastofibrosis, Wallerian-like degeneration in peripheral nerves and deep tissue fibrosis found in the base of the crease.     

γ-Catenin-Dependent Signals Maintain BCR-ABL1+ B Cell Acute Lymphoblastic Leukemia

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Sterilization analysis of contaminated healing abutments and impression copings

This study investigated sterilization of used implant impression copings and healing abutments. Components were analyzed after contamination with Enterococcus foecalis, followed by multiple rounds of sterilization by both steam autoclave and Chemiclave protocols. The authors' results demonstrated that used components showed sterility equal to new components without any visible distortion. These data suggest that component resterilization and reuse may be justified or at least considered in clinical practice. Also, implications for cost savings in the placement of implants are advanced.     

Photodynamic inactivation of pathogenic species <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> and <Emphasis Type="Italic">Candida albicans</Emphasis> with lutetium (III) acetate phthalocyanines and specific light irradiation

Photodynamic inactivation (PDI) is a light-associated therapeutic approach suitable for treatment of local acute infections. The method is based on specific light-activated compound which by specific irradiation and in the presence of molecular oxygen produced molecular singlet oxygen and other reactive oxygen species, all toxic for pathogenic microbial cells. The study presents photodynamic impact of two recently synthesized water-soluble cationic lutetium (III) acetate phthalocyanines (LuPc-5 and LuPc-6) towards two pathogenic strains, namely, the Gram-negative bacterium Pseudomonas aeruginosa and a fungus Candida albicans. The photodynamic effect was evaluated for the cells in suspensions and organized in 48-h developed biofilms. The relatively high levels of uptakes of LuPc-5 and LuPc-6 were determined for fungal cells compared to bacterial cells. The penetration depths and distribution of both LuPcs into microbial biofilms were investigated by means of confocal fluorescence microscopy. The photoinactivation efficiency was studied for a wide concentration range (0.85–30 μM) of LuPc-5 and LuPc-6 at a light dose of 50 J cm^?2 from red light-emitting diode (LED; 665 nm). The PDI study on microbial biofilms showed incomplete photoinactivation (<3 logs) for the used gentle drug-light protocol.     

Nonsurgical Endodontic Treatment of Necrotic Teeth Resolved Apical Lesions on Adjacent Implants with Retrograde/Apical Peri-implantitis: A Case Series with 2-year Follow-up

Retrograde (or apical/periapical) peri-implantitis (RPI) presents with radiographic signs of bone loss at the periapical area of endosteal implants and may also present with clinical signs of abscess formation or a sinus tract traceable to the implant periapex. The lesion may form days up to several years after the initial implant placement. In contrast to marginal peri-implantitis, which has a prevalence of 19.83%, RPI may be underreported because many clinicians are currently not aware of this type of lesion. The etiology, although not fully understood, may be attributed to endodontic infection of an adjacent tooth or residual microorganisms present after the extraction of an infected tooth at the implant site. There are several treatment modalities available for the management of RPI. Nonsurgical root canal treatment may be an option if the implant RPI etiology is suspected to be related to an adjacent endodontically involved tooth. In a previous report, surgical treatment modalities to correct RPI were described. This current case series presents 2 cases of RPI in which nonsurgical treatment of the necrotic adjacent teeth resulted in full radiographic and clinical resolution of the adjacent apical peri-implant lesions with 18-month and 2-year follow-ups, respectively. RPI may be prevented by evaluating the endodontic status of natural teeth adjacent to the implants and by addressing endodontic infections near the implant sites. Certain types of implant RPI may successfully be resolved nonsurgically by addressing adjacent endodontic infections as shown by this case series.     

Derivation of na?ve human embryonic stem cells

The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222–9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.It has become clear with the derivation of mouse epiblast stem cells (mEpiSCs) that pluripotency encompasses more than one stage of development (1, 2). The earlier “naïve” stage represents the preimplantation inner cell mass, typified by mouse ES cells (mESCs), and the “primed,” the postimplantation epiblast, typified by mEpiSCs and human ES cells (hESCs). The challenge in naïve cell maintenance has been protecting cells from differentiation stimuli. This has been achieved in mESCs through exposure to leukemia inhibitory factor (LIF), whereas addition of extracellular signal-regulated kinase (MEK) and glycogen synthase kinase 3 (GSK3) inhibitors (2i) in defined medium allows the cells to attain a homogeneous ground state (3). Defining characteristics of the naïve/ground vs. primed states are shown in Fig. S1A. In humans, the naïve stage has been difficult to capture as a stable in vitro state.There are practical advantages that come with a human naïve state. Among them is ease of trypsin passage and developmental capacity. Whole animals can be generated from good naïve mESCs through tetraploid complementation (4), and mEpiSCs cannot contribute to chimerism. Being more comparable to mESCs, naïve hESCs will likely allow increased developmental potential and a more accurate correlation to mESC data.It has been reported that human induced pluripotent cells (h-iPSCs) can be maintained in the naïve state if the pluripotency-inducing transgenes are not silenced (5). Only recently have hESCs been maintained in a naïve state without transgenes (6). Our primary aim was to generate naïve hESCs not dependent upon transgenes for stable culture. We toggled existing human ESC and mouse mEpiSC lines back from the primed state to grow under the influence of 2i without the need for Activin A. This helped us to define appropriate culture conditions for human naïve cells and allowed the de novo derivation of a naïve hESC line, Elf1. We report on the naïve state of human ESCs capable of unlimited culture in 2i.     

ED2-positive perivascular cells act as neuronophages during delayed neuronal loss in the facial nucleus of the rat

Injection of Fluoro-Gold (FG) into the whisker pad of rats yields a stable retrograde labeling of facial motoneurons. After removal of 10 mm from the facial nerve the microglia phagocytose the FG-prelabeled dead neurons and assume the label. A subsequent brightfield immunostaining of the sections with HRP-DAB as end-product fully quenches the fluorescence of FG from all specifically stained structures (immunoquenching). Combining FG-labeling of neuronophages with immunoquenching, we recently described a population of enigmatic fluorescent cells, found in immediate vicinity to the motoneurons after the general neuronofugal migration of microglia. As the fluorescence of these cells was not quenched after a triple immunostaining with anti neuron-specific enolase, anti-GFAP, and OX-42 (quenching all fluorescence from neurons and glia), they seemed to represent a new, immunologically not identified neuronophage. Now we have further characterized this cell type. Following triple immunostaining, we tested a broad panel of mabs (OX-33, OX-19, OX-18, OX-6, R73, ED1, and ED2) to stain, quench fluorescence, and thus immunotype the unknown phagocytes. Only the mab ED2, the classical marker for perivascular cells, specifically stained the small round neuronophages. This surprising migration of perivascular cells toward decaying neurons was additionally tested and confirmed by intracerebroventricular application of FG prior to resection of the facial nerve Providing evidence for neuronophagia by ED2-positive cells, our results strongly support the hypothesis that the latter are the APC (antigen presenting cells) of the CNS. © 1996 Wiley-Liss, Inc.     

Galectin-3 is upregulated in microglial cells in response to ischemic brain lesions, but not to facial nerve axotomy

We have recently demonstrated that the beta-galactoside-specific lectin galectin-3 is expressed by microglial cells in vitro, but not by normal resting microglia in vivo. In the present study, we have analyzed the expression of galectin-3 by microglia under traumatic conditions in vivo using two experimental rat models which substantially differ in the severity of lesion related to a breakdown of the blood-brain barrier (BBB) and the occurrence of inflammatory processes. These two features are absent after peripheral nerve lesion and present after cerebral ischemia. Here we show that, following facial nerve axotomy under conditions allowing (nerve anastomosis) or not subsequent regeneration (nerve resection), galectin-3 is not expressed by microglia in the corresponding facial nucleus 1-112 days after lesion. Galectin-3 is also absent in microglia at sites of a defective BBB in the normal brain, such as the circumventricular organs. Following experimental ischemia (i.e., permanent occlusion of the middle cerebral artery), in contrast, galectin-3 becomes strongly expressed by activated microglia as early as 48 hours after trauma, as determined by immunohistochemistry and Western blot analysis. Our findings suggest that the expression of galectin-3 by microglia in vivo correlates with the state of microglial activation.     

Hypoglossal and reticular interneurons involved in oro-facial coordination in the rat

Chewing, swallowing, breathing, and vocalization in mammals require precise coordination of tongue movements with concomitant activities of the mimetic muscles. The neuroanatomic basis for this oro-facial coordination is not yet fully understood. After the stereotaxic microinjection of retrograde and anterograde neuronal tracers (biotin-dextran, Fluoro-Ruby, Fluoro-Emerald, and Fluoro-Gold) into the facial and hypoglossal nuclei of the rat, we report here a direct bilateral projection of hypoglossal internuclear interneurons onto facial motoneurons. We also confirm the existence of a small pool of neurons in the dorsal part of the brainstem reticular formation that project ipsilaterally to both facial and hypoglossal nuclei. For precise tracer injections, both motor nuclei were located and identified by the electrical antidromic activation of their constituent motoneurons. Injections of retrograde tracers into the facial nucleus consistently labeled neurons in the hypoglossal nucleus. These neurons prevalently lay in the ipsilateral side, were small in size, and, like classic intrinsic hypoglossal local-circuit interneurons, had several thin dendrites. Reverse experiments - injections of anterograde tracers into the hypoglossal nucleus - labeled fine varicose nerve fiber terminals in the facial nucleus. These fiber terminals were concentrated in the intermediate subdivision of the facial nucleus, with a strong ipsilateral prevalence. Double injections of different tracers into the facial and the hypoglossal nuclei revealed a small, but constant, number of double-labeled neurons located predominantly ipsilateral in the caudal brainstem reticular formation. Hypoglossal internuclear interneurons projecting to the facial nucleus, as well as those neurons of the parvocellular reticular formation that project to both facial and hypoglossal nuclei, could be involved in oro-facial coordination.