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OBJECTIVES: This study aimed to establish a fully digital measurement protocol for standardizing the description of hard palate and cleft morphology in neonates with an isolated cleft palate (CPO) and Pierre Robin sequence (PRS). MATERIALS AND METHODS: A total of 20 digitized plaster models of neonates with CPO and 20 digitized plaster models of neonates with PRS were retrospectively investigated. For the control group, the hard palate was segmented from 21 pre-existing 1.5 T MRI datasets of neonates and exported as an STL file. The digital models were marked with predefined reference points by three raters. Distance, angular, and area measurements were performed using Blender and MeshLab. RESULTS: Neonates with CPO (20.20 ± 2.33 mm) and PRS (21.41 ± 1.81 mm) had a significantly shorter hard palate than the control group (23.44 ± 2.24 mm) (CPO vs. control: P < .001; PRS vs. control: P = .014). Notably, neonates with PRS (33.05 ± 1.95 mm) demonstrated a significantly wider intertuberosity distance than those with CPO (30.52 ± 2.28 mm) (P = .012). Furthermore, there were also significant differences measured between the cleft and control groups (25.22 ± 2.50 mm) (P < .001). CONCLUSIONS: The data from this study demonstrate the feasibility of using MRI datasets to generate digital models of the hard palate. The presence of a cleft palate leads to pronounced adaptations of the total palatal surface area, dorsal width, and length of the hard palate. Mandibular retrognathia and altered tongue position in PRS, as opposed to CPO, might further impact palatal morphology and intertuberosity distance.
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The maxilla occupies a key position in dentofacial orthopaedics, since its transversal development can be directly influenced by orthodontic therapy. The maturation stages of the mid-palatal suture, which are obtained from cone-beam computed tomography images (CBCT), present an addition to clinical decision-making in transversal discrepancies of the upper jaw. In an endeavour to reduce ionizing radiation in adolescents and young adults, who are particularly susceptible to long term stochastic irradiation effects, we investigated the feasibility of 3 Tesla (3T) MRI in detecting the maturation stages of the mid-palatal suture. A collective of 30 patients aged 24-93 years with routine neck MRI at 3T, underwent an additional three-dimensional isotropic T1 weighted study sequence of the midface. Image evaluation was performed on axial, multi-planar formatted reconstructions of the dataset aligned to the midline axis of the palate, and curved reconstructions aligned to the concavity of the palate. Inverted images helped to achieve an image impression similar to the well-known CBCT appearance. All datasets were reviewed by three readers and mid-palatal maturation was scored twice according to Angelieri et al. Intra- and inter-rater agreement were evaluated to measure the robustness of the images for clinical evaluation. 3T MRI deemed reliable for the assessment of mid-palatal suture maturation and hence for the appraisal of the hard palate and its adjacent sutures. The data of this pilot study display the feasibility of non-ionizing cross-sectional MRI for the determination of sutural maturation stages. These findings underline the potential of MRI for orthodontic treatment planning, further contributing to the avoidance of unnecessary radiation doses.
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While the involvement of thermosensitive transient receptor potential channels (TRPs) in dry eye disease (DED) has been known for years, their expression in the meibomian gland (MG) has never been investigated. This study aims to show their expression and involvement in the lipogenesis of the MG, providing a possible new drug target in the treatment of DED. Our RT-PCR, Western blot and immunofluorescence analysis showed the expression of TRPV1, TRPV3, TRPV4 and TRPM8 in the MG at the gene and the protein level. RT-PCR also showed gene expression of TRPV2 but not TRPA1. Calcium imaging and planar patch-clamping performed on an immortalized human meibomian gland epithelial cell line (hMGECs) demonstrated increasing whole-cell currents after the application of capsaicin (TRPV1) or icilin (TRPM8). Decreasing whole-cell currents could be registered after the application of AMG9810 (TRPV1) or AMTB (TRPM8). Oil red O staining on hMGECs showed an increase in lipid expression after TRPV1 activation and a decrease after TRPM8 activation. We conclude that thermo-TRPs are expressed at the gene and the protein level in MGs. Moreover, TRPV1 and TRPM8's functional expression and their contribution to their lipid expression could be demonstrated. Therefore, TRPs are potential drug targets and their clinical relevance in the therapy of meibomian gland dysfunction requires further investigation.
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Disfunção da Glândula Tarsal , Glândulas Tarsais , Humanos , Lipogênese/genética , Western Blotting , Capsaicina/farmacologiaRESUMO
Meibomian gland dysfunction (MGD) is one of the main causes of dry eye disease. To better understand the physiological functions of human meibomian glands (MGs), the present study compared MGs with free sebaceous glands (SGs) and hair-associated SGs of humans using morphological, immunohistochemical, and liquid chromatography-mass spectrometry (LCMS)-based lipidomic approaches. Eyelids with MGs, nostrils, lips, and external auditory canals with free SGs, and scalp with hair-associated SGs of body donors were probed with antibodies against cytokeratins (CK) 1, 8, 10, and 14, stem cell markers keratin 15 and N-cadherin, cell-cell contact markers desmoglein 1 (Dsg1), desmocollin 3 (Dsc3), desmoplakin (Dp), plakoglobin (Pg), and E-cadherin, and the tight junction protein claudin 5. In addition, Oil Red O staining (ORO) was performed in cryosections. Secretions of MGs as well as of SGs of nostrils, external auditory canals, and scalps were collected from healthy volunteers, analyzed by LCMS, and the data were processed using various multivariate statistical analysis approaches. Serial sections of MGs, free SGs, and hair-associated SGs were 3D reconstructed and compared. CK1 was expressed differently in hair-associated SGs than in MGs and other free SGs. The expression levels of CK8, CK10, and CK14 in MGs were different from those in hair-associated SGs and other free SGs. KRT15 was expressed differently in hair-associated SGs, whereas N-cadherin was expressed equally in all types of glands. The cell-cell contact markers Dsg1, Dp, Dsc3, Pg, and E-cadherin revealed no differences. ORO staining showed that lipids in MGs were more highly dispersed and had larger lipid droplets than lipids in other free SGs. Hair-associated SGs had a smaller number of lipid droplets. LCMS revealed that the lipid composition of meibum was distinctively different from that of the sebum of the nostrils, external auditory canals, and scalp. The 3D reconstructions of the different glands revealed different morphologies of the SGs compared with MGs which are by far the largest type of glands. In humans, MGs differ in their morphology and secretory composition and show major differences from free and hair-associated SGs. The composition of meibum differs significantly from that of sebum from free SGs and from hair-associated SGs. Therefore, the MG can be considered as a highly specialized type of holocrine gland that exhibits all the histological characteristics of SGs, but is significantly different from them in terms of morphology and lipid composition.
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Glândulas Tarsais , Glândulas Sebáceas , Humanos , Glândulas Tarsais/metabolismo , Lágrimas/metabolismo , Biomarcadores/metabolismo , Lipídeos/química , Caderinas/metabolismoRESUMO
BACKGROUND: Understanding the attachment patterns of the corrugator supercilii muscle (CS) is vital for treatments designed to improve facial symmetry and functionality. OBJECTIVES: The aim of this research was to elucidate the anatomical trajectory and intricate relationships of the CS within the upper face and midface, specifically focusing on its connections with the frontalis (FT) and orbicularis oculi (OOc) muscles. METHODS: The CS was examined in 41 specimens of embalmed adult Korean cadavers using microdissection, histological analyses, and microcomputed tomography, an in-depth exploration of its anatomical positioning and intricate interactions with adjacent muscles. RESULTS: Some lower fibers of the CS extended to the upper orbital part of the OOc in 59.5% of cases, while the CS interdigitated or blended exclusively with the FT in 40.5% of cases. The fibers of the CS demonstrated diverse extensions towards the upper face and midface, exhibiting varied trajectories and lengths. Additionally, lower fibers of the CS extended to significant anatomical landmarks such as the OOc, malaris muscle, and the superficial musculoaponeurotic system (SMAS). CONCLUSIONS: This study demonstrates that precise understanding of the CS and its relationship with the FT and OOc is crucial for optimizing invasive or non-invasive treatment like botulinum toxin injection, SMAS lifting and browplasty surgery. The extension of lower fibers of the CS to significant anatomical landmarks indicates complex interactions with adjacent facial structures, highlighting the necessity of detailed anatomical knowledge for clinical applications.
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OBJECTIVES: Skull morphology and growth patterns are essential for orthodontic treatment, impacting clinical decision making. We aimed to determine the association of different cephalometric skeletal configurations on midface parameters as measured in 3D CT datasets. MATERIALS AND METHODS: After sample size calculation, a total of 240 fully dentulous patients between 20 and 79 years of age (mean age: 42 ± 15), who had received a CT of the skull within the scope of trauma diagnosis or intracranial bleeding, were retrospectively selected. On the basis of cephalometric analysis, using MPR reconstructions, patients were subdivided into three different vertical skull configurations (brachyfacial, mesofacial, dolichofacial) and the respective skeletal Class I, II, and III relationships. Anatomic parameters were measured using a three-dimensional post-processing console: the thickness of the maxillary and palatine bones as well as the alveolar crest, maxillary body and sutural length, width and height of the hard palate, maxillary facial wall thickness, and masseter muscle thickness and length. RESULTS: Individuals with brachyfacial configurations had a significantly increased palatal and alveolar ridge thicknesses compared to those with dolichofacial- or mesofacial configurations. Brachyfacial configurations presented a significantly increased length and thickness of the masseter muscle (4.599 cm; 1.526 cm) than mesofacial (4.431 cm; 1.466 cm) and dolichofacial configurations (4.405 cm; 1.397 cm) (p < 0.001). Individuals with a skeletal Class III had a significantly shorter palatal length (5.313 cm) than those with Class I (5.406 cm) and Class II (5.404 cm) (p < 0.01). Sutural length was also significantly shorter in Class III (p < 0.05). CONCLUSIONS: Skeletal configurations have an impact on parameters of the bony skull. Also, measurable adaptations of the muscular phenotype could result. CLINICAL RELEVANCE: The association between viscerocranial morphology and midface anatomy might be beneficial for tailoring orthodontic appliances to individual anatomy and planning cortically anchored orthodontic appliances.
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Face , Maxila , Adulto , Humanos , Pessoa de Meia-Idade , Estudos Retrospectivos , Face/anatomia & histologia , Cefalometria/métodos , Maxila/diagnóstico por imagem , Maxila/anatomia & histologia , Palato DuroRESUMO
PURPOSE: To evaluate the feasibility of studying the vascular supply of the orbital and palpebral lobes of the human lacrimal gland using micro-computed tomography (micro-CT) and microscopic dissection. METHODS: The lacrimal gland artery of a fresh parasagittalized cadaver head (male, aged 76 years) was infused with a lead oxide-latex mixture near the occipital pole of the gland. The entire lacrimal gland was imaged using micro-CT and 3D cinematic rendering (CR) and then dissected under a surgical microscope. RESULTS: Micro-CT and CR images showed well-demarcated internal vascular branches of the lacrimal artery and their distribution within the orbital and palpebral lobes. The entire course of the artery and its branches could be visualized by CR and microscopic dissection, with the former showing better spatial orientation and finer branching. The main artery runs along the free edge of the aponeurosis of the levator palpebrae superior muscle and lies in the isthmus portion of the gland (between the orbital and palpebral lobes). The branches of the main lacrimal artery include one branch to the orbital adipose tissue just before entering the gland, two branches to the orbital lobe (medial and lateral), and two branches to the palpebral lobe (medial and lateral). The main artery terminates as palpebral and orbital lobe branches in the lateral half of the lacrimal gland. CONCLUSION: Latex and contrast-enhanced micro-CT is very well suited to visualize the vascular anatomy of the lacrimal artery within the gland. A large number of lacrimal gland examinations using the method presented here are required to demonstrate and understand the variability of the vascular anatomy of the human lacrimal gland.
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Aparelho Lacrimal , Humanos , Masculino , Aparelho Lacrimal/diagnóstico por imagem , Aparelho Lacrimal/anatomia & histologia , Microtomografia por Raio-X , Látex , Artérias/anatomia & histologia , CadáverRESUMO
BACKGROUND: The pectoralis minor muscle is located in the anterior thoracic wall. Typically, is constituted by a single belly originating from the 3rd to the 5th rib and inserted into the coracoid process near the origins of the biceps brachii shorth head and of the coracobrachialis muscle. The current study, on human fetuses, aims to detect all morphological muscle variants and to create a new classification system. MATERIAL AND METHODS: Classical dissection of the thoracic wall and the upper limb was bilaterally performed on 25 (13 male and 12 female) human formalin-fixed fetuses aged 18-38 weeks of gestation. The spontaneously aborted fetuses were donated after parental consent to the Medical University anatomy program. The pectoralis minor muscle's morphology, the number of the muscle's bellies, their origins, and insertions, as well as the morphometric details of each belly of the pectoralis minor, were assessed. RESULTS: The pectoralis minor was bilaterally found in all fetuses (50 cases). Three types of muscle were identified based on the number of muscle bellies. In type, I (typical anatomy), were classified the cases with a single belly (in 66%). This type was divided into two subtypes (Ia and Ib). In the subtype Ia, the single belly had a typical course, and in Ib, a proximal attachment was characterized by two small bellies connecting together and creating one muscular mass. In type II, two bellies (24%), and in type III, three bellies (10%) were identified. CONCLUSIONS: Pectoralis minor is morphologically variable in the number of its bellies, its course, its origins, its insertions, and the location of its proximal attachments. The most common type (typical anatomy) was the type I represented by one belly. Other identified variants in the number of bellies by the present study may be hypothetically a result of prematurely terminated embryogenesis.
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Feto , Músculos Peitorais , Humanos , Músculos Peitorais/anatomia & histologia , Feminino , Feto/anatomia & histologia , Masculino , Dissecação , Idade GestacionalRESUMO
INTRODUCTION: This systematic review aimed to compare reciprocating with continuous rotary instrumentation kinematics, by means of microcomputed tomography evaluations, in extracted human permanent teeth with moderate and severe canal curvatures. METHODS: The research protocol was registered in the International Prospective Register of Systematic Reviews and given the reference number CRD42023404035. An electronic search was undertaken in MEDLINE (PubMed), EBSCO, Scopus, Web of Science databases until December 2021. Manual screening of issues in endodontic journals and references of relevant articles were assessed individually. The risk of bias (RoB) of the included articles was evaluated with the QUIN tool (Quality Assessment Tool for In Vitro Studies). RESULTS: Among 1640 retrieved articles, 49 were included in the qualitative synthesis. Fifteen articles had low RoB, 33 articles had medium RoB, and only 1 study was at high RoB. Continuous rotary systems had better centering ability in both moderate and severe canal curvatures and resulted in less apical transportation in severely curved root canals. None of the kinematic systems was capable of instrumenting the entire canal surface area. Reciprocating kinematics systems tended to provide higher increase in surface area of severely curved canals and produced fewer dentinal microcracks in moderately curved canals. CONCLUSIONS: The evidence presented in this review suggests that continuous rotary system seems to be better than reciprocating system in solving the major issues encountered during root canal instrumentation of extracted teeth with moderate and severe curvatures.
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Cavidade Pulpar , Preparo de Canal Radicular , Microtomografia por Raio-X , Humanos , Fenômenos Biomecânicos , Instrumentos Odontológicos , Cavidade Pulpar/diagnóstico por imagem , Cavidade Pulpar/anatomia & histologia , Desenho de Equipamento , Níquel , Preparo de Canal Radicular/instrumentação , TitânioRESUMO
Voice production has been an area of interest in science since ancient times, and although advancing research has improved our understanding of the anatomy and function of the larynx, there is still little general consensus on these two topics. This review aims to outline the main developments in this field and highlight the areas where further research is needed. The most important hypotheses are presented and discussed highlighting the four main lines of research in the anatomy of the human larynx and their most important findings: (1) the arrangement of the muscle fibers of the thyroarytenoid muscle is not parallel to the vocal folds in the internal part (vocalis muscle), leading to altered properties during contraction; (2) the histological structure of the human vocal cords differs from other striated muscles; (3) there is a specialized type of heavy myosin chains in the larynx; and (4) the neuromuscular system of the larynx has specific structures that form the basis of an intrinsic laryngeal nervous system. These approaches are discussed in the context of current physiological models of vocal fold vibration, and new avenues of investigation are proposed.
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Músculos Laríngeos , Prega Vocal , Voz , Humanos , Músculos Laríngeos/anatomia & histologia , Músculos Laríngeos/fisiologia , Músculos Laríngeos/inervação , Voz/fisiologia , Prega Vocal/anatomia & histologia , Prega Vocal/fisiologia , Laringe/anatomia & histologia , Laringe/fisiologia , AnimaisRESUMO
The fibularis brevis muscle belongs to the lateral compartment of the leg. It originates from the lower two-thirds of the lateral surface of the fibula bone and it also is attached to the anterior intermuscular septum. Its insertion is normally located on the tuberosity at the base of the fifth metatarsal bone, on its lateral side. However, distal attachment of this muscle is morphologically variable. Different variations have been identified and some classification systems have been created for both adults and foetuses. Based on various literature references, we have created a new classification system and compared the frequency of each type. The main aim of this review is to present morphological variations of the fibularis brevis tendon with their clinical significance, especially in autografting in case of surgical treatment of fibularis brevis tendon rupture. The clinical aspect of this pathology and some medical cases will also be presented.
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Fíbula , Músculo Esquelético , Tendões , Humanos , Tendões/anatomia & histologia , Músculo Esquelético/anatomia & histologia , Fíbula/anatomia & histologiaRESUMO
PURPOSE: To elucidate whether it is feasible to use porcine eyes from scalded, abattoir-acquired animals for refractive femtosecond laser research. METHODS: An infrared laser (FS 200) and an ultraviolet laser (prototype version) were tested for their applicability on scalded pig eyes. Fifty porcine eyes were divided into two equally-sized groups and assigned to either the infrared or the ultraviolet laser. Both laser groups were comprised of five subgroups of n = 5 eyes each. Group A: non-scalded eyes (negative control); group B: eyes taken from tunnel-scalded animals; group C1: eyes taken from tank-scalded animals without opaque corneal lesion; group C2: eyes taken from animals with opaque corneal lesion; group D: eyes scalded in toto in the laboratory (positive control). In each group the lasers were employed to create a stromal flap. The quality of the laser cuts and the resulting flap beds, as well as of the porcine corneas themselves, was examined by anterior segment optical coherence tomography and scanning electron microscopy. RESULTS: All scalded specimens exhibited substantial corneal swelling, most pronounced in group C2. After ultraviolet laser application, the tank- and tunnel-scalded samples displayed marked irregularities and an increased degree of surface roughness in the flap beds. After infrared laser application, this was only the case in the tank-scalded specimens. CONCLUSION: It is not recommended to use eyes taken from scalded pigs for ultraviolet femtosecond laser experiments. For infrared femtosecond lasers, eyes taken from tunnel-scalded animals may represent an acceptable alternative, if non-scalded eyes are not available.
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Matadouros , Ceratomileuse Assistida por Excimer Laser In Situ , Suínos , Animais , Ceratomileuse Assistida por Excimer Laser In Situ/métodos , Córnea/patologia , Lasers , Procedimentos Cirúrgicos Oftalmológicos , Lasers de Excimer/uso terapêutico , Substância Própria/cirurgiaRESUMO
The ocular system is in constant interaction with the environment and with numerous pathogens. The ATP-binding cassette (ABC) transporters represent one of the largest groups among the transmembrane proteins. Their relevance has been demonstrated for their defense function against biotic and abiotic stress factors, for metabolic processes in tumors and for their importance in the development of resistance to drugs. The aim of this study was to analyze which ABC transporters are expressed at the ocular surface and in the human lacrimal apparatus. Using RT-PCR, all ABC transporters known to date in humans were examined in tissue samples from human cornea, conjunctiva, meibomian glands and lacrimal glands. The RT-PCR analyses revealed the presence of all ABC transporters in the samples examined, although the results for some of the 48 transporters known in human and analyzed were different in the various tissues. The present results provide information on the expression of ABC transporters at the mRNA level on the ocular surface and in the lacrimal system. Their detection forms the basis for follow-up studies at the protein level, which will provide more information about their physiological significance at the ocular surface and in the lacrimal system and which may explain pathological effects such as drug resistance.
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Transportadores de Cassetes de Ligação de ATP , Túnica Conjuntiva , Córnea , Aparelho Lacrimal , Humanos , Aparelho Lacrimal/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Córnea/metabolismo , Túnica Conjuntiva/metabolismo , Glândulas Tarsais/metabolismo , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Adulto , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
INTRODUCTION: The purpose of this study was to characterize the morphological variations in the quadratus femoris muscle (QF) and to create an anatomical classification that could be used in the planning of surgical procedures in this area, radiological imaging, and rehabilitation. MATERIALS AND METHODS: Ninety-two lower limbs from 46 cadavers, fixed in 10â¯% formalin solution, were examined. RESULTS: The QF muscle was present in all specimens. According to morphology, the QF muscle was classified into three types. The most common type was Type I, characterized by one muscular belly (78.3â¯%), while the second most common type was Type II, characterized by two bellies, was observed in 17.4â¯% of cases. The rarest type was Type III. It was characterized by three bellies and was found in 4.3â¯% of the cases. CONCLUSIONS: The current classification system on quadratus femoris morphological variability is novel. Morphological variants may contribute to clinical issues, such as the ischiofemoral impingement syndrome, that could arise from type I quadratus femoris. Hence, the current study may be applicated to planning surgical procedures, imaging, and rehabilitation.
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Cadáver , Músculo Esquelético , Humanos , Masculino , Feminino , Idoso , Músculo Esquelético/anatomia & histologia , Idoso de 80 Anos ou mais , Pessoa de Meia-IdadeRESUMO
The lacrimal gland is crucial for maintaining ocular health by producing the aqueous component of the tear film, which hydrates and nourishes the ocular surface. Decreased production of this component results in dry eye disease, a condition affecting over 250 million people worldwide. However, the scarcity of primary human material for studying its underlying mechanisms and the absence of a cell model for human lacrimal gland epithelial cells present significant challenges. Here, we describe the generation of immortalized human lacrimal gland cell lines through the introduction of an SV40 antigen. We successfully isolated and characterized three cell clones from a female lacrimal gland donor, confirming their epithelial identity through genomic and protein analyses, including PCR, RNAseq, immunofluorescence and cultivation in a 3D spheroid model. Our findings represent a significant advancement, providing improved accessibility to investigate the molecular pathogenesis mechanisms of dry eye disease and potential therapeutic interventions. We identified the expression of typical epithelial cell marker genes and demonstrated the cells' capability to form 2D cell sheets and 3D spheroids. This establishment of immortalized human lacrimal gland cells with epithelial characteristics holds promise for future comprehensive studies, contributing to a deeper understanding of dry eye disease and its cellular mechanisms.
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Síndromes do Olho Seco , Aparelho Lacrimal , Humanos , Feminino , Aparelho Lacrimal/metabolismo , Lágrimas/metabolismo , Síndromes do Olho Seco/metabolismo , Linhagem CelularRESUMO
Although 2D cancer models have been the standard for drug development, they don't resemble in vivo properties adequately. 3D models can potentially overcome this. Bioprinting is a promising technique for more refined models to investigate central processes in tumor development such as proliferation, dormancy or metastasis. We aimed to analyze bioinks, which could mimic these different tumor stages in a cast vascularized arteriovenous loop melanoma model in vivo. It has the advantage to be a closed system with a defined microenvironment, supplied only with one vessel-ideal for metastasis research. Tested bioinks showed significant differences in composition, printability, stiffness and microscopic pore structure, which led to different tumor stages (Matrigel and Alg/HA/Gel for progression, Cellink Bioink for dormancy) and resulted in different primary tumor growth (Matrigel significantly higher than Cellink Bioink). Light-sheet fluorescence microscopy revealed differences in vascularization and hemorrhages with no additional vessels found in Cellink Bioink. Histologically, typical human melanoma with different stages was demonstrated. HMB-45-positive tumors in progression inks were infiltrated by macrophages (CD163), highly proliferative (Ki67) and metastatic (MITF/BRN2, ATX, MMP3). Stainings of lymph nodes revealed metastases even without significant primary tumor growth in Cellink Bioink. This model can be used to study tumor pathology and metastasis of different tumor stages and therapies.
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Micro- and nanoplastics (MNPs) are ubiquitous in the environment, resulting in the uptake of MNPs by a variety of organisms, including humans, leading to particle-cell interaction. Human macrophages derived from THP-1 cell lines take up Polystyrene (PS), a widespread plastic. The question therefore arises whether primary human macrophages also take up PS micro- and nanobeads (MNBs) and how they react to this stimulation. Major aim of this study is to visualize this uptake and to validate the isolation of macrophages from peripheral blood mononuclear cells (PBMCs) to assess the impact of MNPs on human macrophages. Uptake of macrophages from THP-1 cell lines and PBMCs was examined by transmission electron microscopy (TEM), scanning electron microscopy and live cell imaging. In addition, the reaction of the macrophages was analyzed in terms of metabolic activity, cytotoxicity, production of reactive oxygen species (ROS) and macrophage polarization. This study is the first to visualize PS MNBs in primary human cells using TEM and live cell imaging. Metabolic activity was size- and concentration-dependent, necrosis and ROS were increased. The methods demonstrated in this study outline an approach to assess the influence of MNP exposure on human macrophages and help investigating the consequences of worldwide plastic pollution.
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Macrófagos , Microplásticos , Poliestirenos , Espécies Reativas de Oxigênio , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Poliestirenos/química , Poliestirenos/toxicidade , Células THP-1 , Microplásticos/toxicidade , Leucócitos Mononucleares/efeitos dos fármacos , Nanopartículas/toxicidade , Nanopartículas/química , Sobrevivência Celular/efeitos dos fármacos , Microscopia Eletrônica de Transmissão , Tamanho da PartículaRESUMO
PURPOSE: Diabetes mellitus (DM) is a leading risk factor for corneal neuropathy and dry eye disease (DED). Another common consequence of DM is diabetic peripheral polyneuropathy (DPN). Both complications affect around 50 % of the DM patients but the relationship between DM, DED and DPN remains unclear. METHODS: In this study, we examined mice with early onset of DM and PN after streptozotocin (STZ)-induced diabetes (DPN). We compared the early morphological changes of the sciatic nerve, dorsal root and trigeminal ganglia with the changes in the ocular surface, including tear proteomic and we also investigated respective changes in the gene expressions and morphological alterations in the eye tissues involved in tear production. RESULTS: The lacrimal gland, conjunctival goblet cells and cornea showed morphological changes along with alterations in tear proteins without any obvious signs of ocular surface inflammation. The gene expression for respectively altered tear proteins i.e., of Clusterin in cornea, Car6, Adh3a1, and Eef1a1 in eyelids, and Pigr in the lacrimal gland also showed significant changes compared to control mice. In the trigeminal ganglia like in the dorsal root ganglia neuronal cells showed swollen mitochondria and, in the latter, there was a significant increase of NADPH oxidases and MMP9 suggestive of oxidative and neuronal stress. In the dorsal root ganglia and the sciatic nerve, there was an upregulation of a number of pro-inflammatory cytokines and pain-mediating chemokines. CONCLUSION: The early ocular changes in DM Mice only affect the lacrimal gland. Which, is reflected in the tear film composition of DPN mice. Due to the high protein concentration in tear fluid in humans, proteomic analysis in addition to noninvasive investigation of goblet cells and cornea can serve as a tools for the early diagnosis of DPN, DED in clinical practice. Early treatment could delay or even prevent the ocular complications of DM such as DED and PN.
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Diabetes Mellitus , Neuropatias Diabéticas , Síndromes do Olho Seco , Aparelho Lacrimal , Humanos , Camundongos , Animais , Estreptozocina/metabolismo , Neuropatias Diabéticas/metabolismo , Proteômica , Aparelho Lacrimal/metabolismo , Lágrimas/metabolismo , Síndromes do Olho Seco/diagnóstico , Inflamação/metabolismoRESUMO
OBJECTIVE: Perineuronal nets (PNN) are specialized extracellular matrix (ECM) components of the central nervous system, frequently accumulating at the surface of inhibitory GABAergic interneurons. While an altered distribution of PNN has been observed in neurological disorders including Alzheimer's disease, schizophrenia and epilepsy, their anatomical distribution also changes during physiological brain maturation and aging. Such an age-dependent shift was experimentally associated also with hippocampal engram formation during brain maturation. Our aim was to histopathologically assess PNN in the hippocampus of adult and pediatric patients with temporal lobe epilepsy (TLE) compared to age-matched post-mortem control subjects and to compare PNN-related changes with memory impairment observed in our patient cohort. METHODS: Sixty-six formalin-fixed and paraffin-embedded tissue specimens of the human hippocampus were retrieved from the European Epilepsy Brain Bank. Twenty-nine patients had histopathologically confirmed hippocampal sclerosis (HS), and eleven patients suffered from TLE without HS. PNN were immunohistochemically visualized using an antibody directed against aggrecan and manually counted from hippocampus subfields and the subiculum. RESULTS: PNN density increased with age in both human controls and TLE patients. However, their density was significantly higher in all HS patients compared to age-matched controls. Intriguingly, TLE patients presented presurgically with better memory when their hippocampal PNN density was higher (p < 0.05). SIGNIFICANCE: Our results were compatible with age-dependent ECM specialization in the human hippocampus and its precocious aging in the epileptic condition. These observations confirm recent experimental animal models and also support the notion that PNN play a role in memory formation in the human brain. PLAIN LANGUAGE SUMMARY: "Perineuronal nets" (PNN) are a specialized compartment of the extracellular matrix (ECM), especially surrounding highly active neurons of the mammalian brain. There is evidence that PNN play a role in memory formation, brain maturation, and in some pathologies like Alzheimer's disease, schizophrenia or epilepsy. In this study, we investigated the role of PNN in patients suffering from drug-resistant focal epilepsy compared to controls. We found that with increasing age, more neurons are surrounded by PNN. Similarly, all epilepsy patients but especially patients with better memory performance also had more PNN. This study raises further interest in studying ECM molecules in the human brain under physiological and pathophysiological conditions.