Department of Molecular Cell Biology, University of Maastricht, Research Institutes CARIM, GROW, and EURON, Maastricht; Department of Biomechanics and Tissue Engineering, Faculty of Biomedical ...Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands; Institut National de la Santé et de la Recherche Médicale U.582; Université Pierre et Marie Curie-Paris 6, Faculté de Médecine; AP-HP, Groupe Hospitalier Pitié-Salpêtrière, U. F. Myogénétique et Cardiogénétique, Service de Biochimie and Association Institut de Myologie, Groupe Hospitalier Pitié-Salpêtrière, Paris, France; Integrative Cell Biology Laboratory, School of Biological and Biomedical Sciences, University of Durham, Durham, United Kingdom
It has been demonstrated that nuclear lamins are important proteins in maintaining cellular as well as nuclear integrity, and in maintaining chromatin organization in the nucleus. Moreover, there is growing evidence that lamins play a prominent role in transcriptional control. The family of laminopathies is a fast-growing group of diseases caused by abnormalities in the structure or processing of the lamin A/C ( LMNA) gene. Mutations or incorrect processing cause more than a dozen different inherited diseases, ranging from striated muscular diseases, via fat- and peripheral nerve cell diseases, to progeria. This broad spectrum of diseases can only be explained if the responsible A-type lamin proteins perform multiple functions in normal cells. This review gives an overview of current knowledge on lamin structure and function and all known diseases associated with LMNA abnormalities. Based on the knowledge of the different functions of A-type lamins and associated proteins, explanations for the observed phenotypes are postulated. It is concluded that lamins seem to be key players in, among others, controlling the process of cellular ageing, since disturbance in lamin protein structure gives rise to several forms of premature ageing.
Infection with high-risk human papillomavirus (HPV) type 16 is an independent risk factor for the development of oropharyngeal squamous cell carcinomas (OSCC). However, it is unclear whether viral ...integration is an essential hallmark in the carcinogenic process of OSCC and whether HPV integration correlates with the level of viral gene transcription and influences the expression of disrupted host genes. We analyzed 75 patients with OSCC. HPV16-positivity was proven by p16(INK4A) immunohistochemistry, PCR and FISH. Viral integration was examined using DIPS- as well as APOT-PCR. Viral E2, E6 and E7 gene expression levels were quantified by quantitative reverse transcriptase (RT-q)PCR. Expression levels of 7 human genes disrupted by the virus were extracted from mRNA expression profiling data of 32 OSCCs. Viral copy numbers were assessed by qPCR in 73 tumors. We identified 37 HPV16-human fusion products indicating viral integration in 29 (39%) OSCC. In the remaining tumors (61%) only episome-derived PCR products were detected. When comparing OSCC with or without an integration-derived fusion product, we did not find significant differences in the mean RNA expression of viral genes E2, E6 and E7 or the viral copy numbers per cell, nor did the RNA expression of the HPV-disrupted genes differ from either group of OSCC. In conclusion, our data do not support the hypothesis that integration affects the levels of viral and/or HPV-disrupted human gene transcripts. Thus constitutive, rather than a high level, of expression of oncogene transcripts appears to be required in HPV-related OSCC.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Both apolipoprotein E (apoE) and 24(S)-hydroxycholesterol are involved in the pathogenesis of Alzheimer disease (AD). It has been hypothesized that apoE affects AD development via isoform-specific ...effects on lipid trafficking between astrocytes and neurons. However, the regulation of the cholesterol supply of neurons via apoE-containing high density lipoproteins remains to be clarified. We show for the first time that the brain-specific metabolite of cholesterol produced by neurons, i.e. 24(S)-hydroxycholesterol, induces apoE transcription, protein synthesis, and secretion in a dose- and time-dependent manner in cells of astrocytic but not of neuronal origin. Moreover, 24(S)-hydroxycholesterol primes astrocytoma, but not neuroblastoma cells, to mediate cholesterol efflux to apoE. Similar results were obtained using the synthetic liver X receptor (LXR) agonist GW683965A, suggesting involvement of an LXR-controlled signaling pathway. A 10-20-fold higher basal LXRα and -β expression level in astrocytoma compared with neuroblastoma cells may underlie these differential effects. Furthermore, apoE-mediated cholesterol efflux from astrocytoma cells may be controlled by the ATP binding cassette transporters ABCA1 and ABCG1, since their expression was also up-regulated by both compounds. In contrast, ABCG4 seems not to be involved, because its expression was induced only in neuronal cells. The expression of sterol regulatory element-binding protein (SREBP-2), low density lipoprotein receptor, 3-hydroxy-3-methylglutaryl-CoA reductase, and SREBP-1c was transiently up-regulated by GW683965A in astrocytes but down-regulated by 24(S)-hydroxycholesterol, suggesting that cholesterol efflux and synthesis are regulated independently. In conclusion, evidence is provided that 24(S)-hydroxycholesterol induces apoE-mediated efflux of cholesterol in astrocytes via an LXR-controlled pathway, which may be relevant for chronic and acute neurological diseases.
Different studies have shown that HPV16‐positive OPSCC can be subdivided based on integration status (integrated, episomal and mixed forms). Because we showed that integration neither affects the ...levels of viral genes, nor those of virally disrupted human genes, a genome‐wide screen was performed to identify human genes which expression is influenced by viral integration and have clinical relevance. Thirty‐three fresh‐frozen HPV‐16 positive OPSCC samples with known integration status were analyzed by mRNA expression profiling. Among the genes of interest, Aldo‐keto‐reductases 1C1 and 1C3 (AKR1C1, AKR1C3) were upregulated in tumors with viral integration. Additionally, 141 OPSCC, including 48 HPV‐positive cases, were used to validate protein expression by immunohistochemistry. Results were correlated with clinical and histopathological data. Non‐hierarchical clustering resulted in two main groups differing in mRNA expression patterns, which interestingly corresponded with viral integration status. In OPSCC with integrated viral DNA, often metabolic pathways were deregulated with frequent upregulation of AKR1C1 and AKR1C3 transcripts. Survival analysis of 141 additionally immunostained OPSCC showed unfavorable survival rates for tumors with upregulation of AKR1C1 or AKR1C3 (both p <0.0001), both in HPV‐positive (p ≤0.001) and ‐negative (p ≤0.017) tumors. OPSCC with integrated HPV16 show upregulation of AKR1C1 and AKR1C3 expression, which strongly correlates with poor survival rates. Also in HPV‐negative tumors, upregulation of these proteins correlates with unfavorable outcome. Deregulated AKR1C expression has also been observed in other tumors, making these genes promising candidates to indicate prognosis. In addition, the availability of inhibitors of these gene products may be utilized for drug treatment.
What's new?
Oropharyngeal squamous cell carcinoma (OPSCC) is often associated with high‐risk human papillomavirus infection, especially HPV16, but the clinical relevance of viral integration in a subset of these tumors remains unclear. Here the authors describe transcriptional signatures of metabolic reprogramming including increased expression of the aldo‐keto‐reductases AKR1C1 and AKR1C3 proteins in OPSCC cells regardless of HPV status. Tumors with upregulation of these proteins were strongly associated with poor prognosis suggesting prognostic as well as therapeutic implications for OPSCC patients beyond HPV16 infection.
Pulmonary carcinoids are neuroendocrine tumors histopathologically subclassified into typical (TC; no necrosis, <2 mitoses per 2 mm) and atypical (AC; necrosis or 2 to 10 mitoses per 2 mm). The ...reproducibility of lung carcinoid classification, however, has not been extensively studied and may be hampered by the presence of pyknotic apoptosis mimicking mitotic figures. Furthermore, prediction of prognosis based on histopathology varies, especially for ACs. We examined the presence of interobserver variation between 5 experienced pulmonary pathologists who reviewed 123 originally diagnosed pulmonary carcinoid cases. The tumors were subsequently redistributed over 3 groupsunanimously classified cases, consensus cases (4/5 pathologists rendered identical diagnosis), and disagreement cases (divergent diagnosis by ≥2 assessors). κ-values were calculated, and results were correlated with clinical follow-up and molecular data. When focusing on the 114/123 cases unanimously classified as pulmonary carcinoids, the interobserver agreement was only fair (κ=0.32). Of these 114 cases, 55% were unanimously classified, 25% reached consensus classification, and for 19% there was no consensus. ACs were significantly more often in the latter category (P=0.00038). The designation of TCs and ACs by ≥3 assessors was not associated with prognosis (P=0.11). However, when disagreement cases were allocated on the basis of Ki-67 proliferative index (<5%; ≥5%) or nuclear orthopedia homeobox immunostaining (+; −), correlation with prognosis improved significantly (P=0.00040 and 0.0024, respectively). In conclusion, there is a considerable interobserver variation in the histopathologic classification of lung carcinoids, in particular concerning ACs. Additional immunomarkers such as Ki-67 or orthopedia homeobox may improve classification and prediction of prognosis.
The envelope that encapsulates the cell nucleus has recently gained considerable interest, as several clinical syndromes are linked to mutations in its molecular components. Most disorders recognized ...so far are caused by defects in the nuclear lamins, building blocks of a filamentous network lining the nucleoplasmic side of the inner nuclear membrane. Nuclear lamins are the evolutionary precursors of cytoskeletal intermediate filaments and associate in a head-to-tail manner into a stable lamina at the nuclear periphery and into a more dispersed structure in the nucleoplasm. Lamins have a scaffolding function for several nuclear processes such as transcription, chromatin organization and DNA replication, and maintain nuclear and cellular integrity. Mutations in the LMNA gene, encoding A-type lamins, can cause cardiac and skeletal muscle disease, lipodystrophy and premature ageing phenotypes. Hence, the integrity of the nuclear envelope seems essential for longevity. Furthermore, the laminopathies provide evidence that metabolism and ageing are as tightly linked in humans as they are in model organisms such as C. elegans. In this review, we elaborate on the structure and functions of nuclear lamins, the spectrum of syndromes related to mutations in nuclear envelope components and pathogenic concepts unifying these disorders.
Prolonged atrial fibrillation (AF) results in electrical, structural, and gap-junctional remodeling. We examined the reversibility of the changes in (ultra)structure and gap junctions.
Four groups of ...goats were used: (1) sinus rhythm (SR), (2) 4 months' AF (4 mo AF), (3) 2 months' SR after 4 mo AF (2 mo post-AF), and (4) 4 months' SR after 4 mo AF (4 mo post-AF). Atria were characterized electrophysiologically, (ultra)structure was studied by light and electron microscopy, and structural and gap-junctional protein expression was studied by immunohistochemistry or Western blotting. The atrial effective refractory period had completely returned to normal values 2 mo post-AF. Induced AF episodes still lasted for minutes at 2 and 4 mo post-AF, compared with seconds in the SR group. Structural abnormalities were still present at 2 and 4 mo post-AF, although to a lesser extent. The increased atrial myocyte diameter was back to normal at 4 mo post-AF. The number of myocytes with severe myolysis had almost normalized 4 mo post-AF, whereas myocytes with mild myolysis remained significantly increased. Extracellular matrix area fraction after 4 mo AF was similar to SR. However, the extracellular matrix fraction per myocyte had increased after 4 mo AF and remained higher post-AF. Changes in expression of structural proteins were partially restored post-AF. The reduction of connexin 40 that was observed during AF was completely reversed at 4 mo post-AF.
Recovery from structural remodeling after 4 mo AF is a slow process and is still incomplete 4 mo post-AF. Several months post-AF, the duration of AF episodes is still prolonged (minutes).
The response of individual cells to cellular stress is vital for cellular functioning. A large network of physically interconnected cellular components, starting from the structural components of the ...cells' nucleus, via cytoskeleton filaments to adhesion molecules and the extracellular matrix, constitutes an integrated matrix that functions as a scaffold allowing the cell to cope with mechanical stress. Next to a role in mechanical properties, this network also has a mechanotransductional function in the response to mechanical stress. This signaling route does not only regulate a rapid reorganization of structural components such as actin filaments, but also stimulates for example gene activation via NFκB and other transcription factors. The importance of an intact mechano-signaling network is illustrated by the physiological consequences of several genetic defects of cellular network components e.g. actin, dystrophin, desmin and lamins. These give rise to an impaired response of the affected cells to mechanical stress and often result in dystrophy of the affected tissue. Recently, the importance of the cell nucleus in cellular strength has been established. Several new interconnecting proteins, such as the nesprins that link the nuclear lamina to the cytoskeleton, have been identified. Furthermore, the function of nuclear lamins in determining cellular strength and nuclear stability was illustrated in lamin-knock-out cells. Absence of the A-type lamins or mutations in these structural components of the nuclear lamina lead to an impaired cellular response to mechanical stress and disturbances in cytoskeletal organization. In addition, laminopathies show clinical phenotypes comparable to those seen for diseases resulting from genetic defects in cytoskeletal components, further indicating that lamins play a central role in maintaining the mechanical properties of the cell.
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