Ana Carolina Prado Ribeiro

Interpretation of the sonic hedgehog morphogen gradient by a temporal adaptation mechanism

Morphogens act in developing tissues to control the spatial arrangement of cellular differentiation. The activity of a morphogen has generally been viewed as a concentration-dependent response to a diffusible signal, but the duration of morphogen signalling can also affect cellular responses. One such example is the morphogen sonic hedgehog (SHH). In the vertebrate central nervous system and limbs, the pattern of cellular differentiation is controlled by both the amount and the time of SHH exposure. How these two parameters are interpreted at a cellular level has been unclear. Here we provide evidence that changing the concentration or duration of SHH has an equivalent effect on intracellular signalling. Chick neural cells convert different concentrations of SHH into time-limited periods of signal transduction, such that signal duration is proportional to SHH concentration. This depends on the gradual desensitization of cells to ongoing SHH exposure, mediated by the SHH-dependent upregulation of patched 1 (PTC1), a ligand-binding inhibitor of SHH signalling. Thus, in addition to its role in shaping the SHH gradient, PTC1 participates cell autonomously in gradient sensing. Together, the data reveal a novel strategy for morphogen interpretation, in which the temporal adaptation of cells to a morphogen integrates the concentration and duration of a signal to control differential gene expression.

Gene Regulatory Logic for Reading the Sonic Hedgehog Signaling Gradient in the Vertebrate Neural Tube

Summary Secreted signals, known as morphogens, provide the positional information that organizes gene expression and cellular differentiation in many developing tissues. In the vertebrate neural tube, Sonic Hedgehog (Shh) acts as a morphogen to control the pattern of neuronal subtype specification. Using an in vivo reporter of Shh signaling, mouse genetics, and systems modeling, we show that a spatially and temporally changing gradient of Shh signaling is interpreted by the regulatory logic of a downstream transcriptional network. The design of the network, which links three transcription factors to Shh signaling, is responsible for differential spatial and temporal gene expression. In addition, the network renders cells insensitive to fluctuations in signaling and confers hysteresis—memory of the signal. Our findings reveal that morphogen interpretation is an emergent property of the architecture of a transcriptional network that provides robustness and reliability to tissue patterning.

Coordination of progenitor specification and growth in mouse and chick spinal cord

Introduction The hallmarks of development are tissue growth and the generation of cell diversity, resulting in reproducibly patterned animals. Yet, how growth and cell fate specification are coordinated to determine the variety of cell types and the proportions of their populations is not well understood. The specification of cell types can be controlled by long-range signals, called morphogens. In some tissues, the shape of morphogen gradients is controlled to match the rate at which the tissue grows, which may keep the pattern proportional to the overall tissue size. To understand whether a similar strategy applies to the patterning of the spinal cord, we measured growth and cell fate specification in chick and mouse embryos of normal size, as well as mutant mice of smaller size. Growth and patterning of the spinal cord. Progenitors are organized in a striped pattern, revealed by their gene expression. In animals of different size, the pattern changes in the same way as the spinal cord grows. This is controlled sequentially, first by progenitor specification by opposing morphogen gradients (bottom left), then by cell-type–specific differentiation resulting in differently shaped clones (green, bottom right). Rationale Morphogen gradients emanating from the dorsal and ventral sides of the spinal cord establish a striped pattern of gene expression in neural progenitors along the dorsoventral axis. Ventrally, Sonic Hedgehog (Shh) is the key morphogen. As the tissue grows in size, the levels of Shh activity in progenitors decrease and are not constant at progenitor domain boundaries. This prompted us to examine whether growth contributed to pattern formation. To this end, we measured the four parameters that control the number of progenitors in a domain: (i) cell proliferation, (ii) cell death, (iii) terminal differentiation of progenitors into postmitotic neurons, and (iv) switches in cell identity—morphogen-driven changes in gene expression respecifying one progenitor type into another. Results Unlike systems in which pattern is proportional to size, we found that the relative dorsoventral sizes of neural progenitor domains change continuously during development . These changes in proportions are conserved between mouse, chick, and smaller mouse mutants. Because the proliferation rate of progenitors was spatially uniform and cell death was negligible, neither of these processes could account for the dynamics of pattern formation. Instead, the data revealed two distinct phases of spinal cord development. Initially, the influence of the morphogens dominates, and switches in cell identity establish pattern. During the second phase, domain-specific differentiation rates emerge, causing changes in the relative proportions of progenitor cell populations. Clonal analysis indicated that this effect is anisotropic: Differentiation affects dorsoventral and apicobasal, but not anterioposterior, domain growth. The outcome is that different domains grow in register along the anterioposterior axis. Consistent with the two-phase model, the switches in cell fate and the sensitivity to changes in morphogen signaling decrease as development proceeds. Conversely, experimentally flattening the difference in differentiation rate between domains during the second phase alters pattern. Conclusion The data reveal two phases of neural tube development and show that sequential control of progenitor cell specification and differentiation elaborates pattern without requiring signaling gradients to expand as tissues grow. Control of the differentiation rate is likely to contribute to pattern formation in other tissues. Furthermore, the domain-specific regulation of the differentiation rates could suggest a means to achieve reproducible development despite differences in individual size. Differentiation rates regulate pool sizes Even though a basketball player is bigger than a gymnast, their neural tubes are organized in the same way. Studying chick and mouse embryos, Kicheva et al. show that rates of cell differentiation are key (see the Perspective by Pourquie). In a two-phase process, signaling sweeps through the neural tube early on to establish some aspects of cell fate, but later, pools of progenitor cells take on their own regulation. A progenitor that differentiates is no longer a progenitor, and thus the rate of differentiation determines the size of the progenitor pool. The relative sizes of progenitor pools shift as development progresses, to build the spinal cord so that everyone, large or small, has the right proportion of each component. Science, this issue p. 10.1126/science.1254927; see also p. 1565 Embryonic chick and mouse reveal how neural tube differentiation takes on new strategies as development progresses. [Also see Perspective by Pourquie] Development requires tissue growth as well as cell diversification. To address how these processes are coordinated, we analyzed the development of molecularly distinct domains of neural progenitors in the mouse and chick neural tube. We show that during development, these domains undergo changes in size that do not scale with changes in overall tissue size. Our data show that domain proportions are first established by opposing morphogen gradients and subsequently controlled by domain-specific regulation of differentiation rate but not differences in proliferation rate. Regulation of differentiation rate is key to maintaining domain proportions while accommodating both intra- and interspecies variations in size. Thus, the sequential control of progenitor specification and differentiation elaborates pattern without requiring that signaling gradients grow as tissues expand.

Ptch1 and Gli regulate Shh signalling dynamics via multiple mechanisms.

In the vertebrate neural tube, the morphogen Sonic Hedgehog (Shh) establishes a characteristic pattern of gene expression. Here we quantify the Shh gradient in the developing mouse neural tube and show that while the amplitude of the gradient increases over time, the activity of the pathway transcriptional effectors, Gli proteins, initially increases but later decreases. Computational analysis of the pathway suggests three mechanisms that could contribute to this adaptation: transcriptional upregulation of the inhibitory receptor Ptch1, transcriptional downregulation of Gli and the differential stability of active and inactive Gli isoforms. Consistent with this, Gli2 protein expression is downregulated during neural tube patterning and adaptation continues when the pathway is stimulated downstream of Ptch1. Moreover, the Shh-induced upregulation of Gli2 transcription prevents Gli activity levels from adapting in a different cell type, NIH3T3 fibroblasts, despite the upregulation of Ptch1. Multiple mechanisms therefore contribute to the intracellular dynamics of Shh signalling, resulting in different signalling dynamics in different cell types.

Osteonecrosis of the mandible associated with bevacizumab therapy

Bevacizumab is a humanized antibody that blocks vascular endothelial growth factor and is of great value for the treatment of advanced cancer. Several adverse effects following its administration have been reported. To date, only 8 cases of osteonecrosis of the jaws associated with bevacizumab (without any association with bisphosphonates) have been reported. The aim of this article was to describe an original case of bevacizumab-related osteonecrosis of the jaw. A 61-year-old man diagnosed with advanced renal cell carcinoma was undergoing treatment with intravenous bevacizumab and temsirolimus when he spontaneously developed mandible osteonecrosis, which resolved after 3 months of conservative management. The present case reinforces recent speculation that the anti-angiogenic properties of bevacizumab may represent a potential new source of osteonecrosis of the jaws in patients undergoing cancer treatment. Multidisciplinary teams in cancer care should be aware of the possible association between osteonecrosis of the jaw and bevacizumab therapy.

High incidences of DNA ploidy abnormalities in tongue squamous cell carcinoma of young patients: an international collaborative study

Santos‐Silva A R, Ribeiro A C P, Soubhia A M P, Miyahara G I, Carlos R, Speight P M, Hunter K D, Torres‐Rendon A, Vargas P A & Lopes M A (2011) Histopathology 58, 1127–1135
High incidences of DNA ploidy abnormalities in tongue squamous cell carcinoma of young patients: an international collaborative study

Bilateral central ossifying fibroma affecting the mandible: report of an uncommon case and critical review of the literature

Ossifying fibroma (OF) is a well demarcated benign neoplasm primarily found in the jaw and composed of fibrocellular tissue and mineralized material. Occurrence of multiple OFs (synchronous) is rare in the jaws, and only 10 cases have been documented. The aim of this report was to present an additional case of bilateral central OF in the mandible of a patient not affected by the hyperparathyroidism–jaw tumors syndrome (HPTJT), emphasizing the features that distinguish this lesion from HPT-JT and performing a critical review of the current literature and concepts. Benign fibro-osseous lesions (FOLs) are a poorly defined and to some extent controversial group of lesions affecting the jaws and craniofacial bones. FOL refers to a group of pathologic processes in which normal bone is replaced by fibroblasts and collagen fibers containing variable amounts of mineralized material. This group encompasses fibrous dysplasia, benign fibro-osseous neoplasms (central ossifying fibroma), and a heterogeneous group of reactive lesions (osseous dysplasias). Because of the histopathologic similarities among these lesions, the definitive diagnosis requires a precise correlation of the clinical, histopathologic, and imaging findings.

Insights into immune responses in oral cancer through proteomic analysis of saliva and salivary extracellular vesicles.

The development and progression of oral cavity squamous cell carcinoma (OSCC) involves complex cellular mechanisms that contribute to the low five-year survival rate of approximately 20% among diagnosed patients. However, the biological processes essential to tumor progression are not completely understood. Therefore, detecting alterations in the salivary proteome may assist in elucidating the cellular mechanisms modulated in OSCC and improve the clinical prognosis of the disease. The proteome of whole saliva and salivary extracellular vesicles (EVs) from patients with OSCC and healthy individuals were analyzed by LC-MS/MS and label-free protein quantification. Proteome data analysis was performed using statistical, machine learning and feature selection methods with additional functional annotation. Biological processes related to immune responses, peptidase inhibitor activity, iron coordination and protease binding were overrepresented in the group of differentially expressed proteins. Proteins related to the inflammatory system, transport of metals and cellular growth and proliferation were identified in the proteome of salivary EVs. The proteomics data were robust and could classify OSCC with 90% accuracy. The saliva proteome analysis revealed that immune processes are related to the presence of OSCC and indicate that proteomics data can contribute to determining OSCC prognosis.

A targeted proteomic strategy for the measurement of oral cancer candidate biomarkers in human saliva

Head and neck cancers, including oral squamous cell carcinoma (OSCC), are the sixth most common malignancy in the world and are characterized by poor prognosis and a low survival rate. Saliva is oral fluid with intimate contact with OSCC. Besides non‐invasive, simple, and rapid to collect, saliva is a potential source of biomarkers. In this study, we build an SRM assay that targets fourteen OSCC candidate biomarker proteins, which were evaluated in a set of clinically‐derived saliva samples. Using Skyline software package, we demonstrated a statistically significant higher abundance of the C1R, LCN2, SLPI, FAM49B, TAGLN2, CFB, C3, C4B, LRG1, SERPINA1 candidate biomarkers in the saliva of OSCC patients. Furthermore, our study also demonstrated that CFB, C3, C4B, SERPINA1 and LRG1 are associated with the risk of developing OSCC. Overall, this study successfully used targeted proteomics to measure in saliva a panel of biomarker candidates for OSCC.

Head and neck amyloidosis: clinicopathological features and immunohistochemical analysis of 14 cases

BACKGROUND Amyloidosis is associated with or caused by amyloid deposition. These fibrillar proteins may be deposited extracellularly causing tissue damage or impairment. OBJECTIVES The aim of the study was to retrospectively review pathology archives in two oral diagnostic centers for cases fulfilling criteria of amyloidosis and to differentiate AA and AL types of amyloidosis. METHODS The clinicopathological features, alkaline Congo red staining, with and without pretreatment with potassium permanganate, and immunohistochemical (IHC) staining with anti-AA, anti-kappa (κ), and anti-lambda (λ) light chain antibodies were carried out and analyzed. RESULTS The search identified 14 cases. Ten patients were women and four were men, with a mean age of 58 years. Eleven patients had systemic involvement by amyloidosis (associated either with multiple myeloma or plasma cell dyscrasia/monoclonal gammopathies), while three presented the localized type, one of them associated with plasmacytoma. All cases showed positivity for κ or λ light chains (AL-amyloid) and presented resistance to the potassium permanganate pretreatment. CONCLUSIONS Our results show that the head and neck region is preferentially affected by systemic AL-amyloidosis, usually associated with plasma cell dyscrasia. Interestingly, two cases affected by inflammatory rheumatic diseases presented AL-amyloid deposition. Moreover, even after pretreatment with potassium permanganate, which was helpful in highlighting the presence of AL-amyloid, in agreement with the IHC findings, clinical classifications should be carefully made in systemic amyloidosis.