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Productive Infection of Human Skeletal Muscle Cells by Pandemic and Seasonal Influenza A(H1N1) Viruses: e79628
Besides the classical respiratory and systemic symptoms, unusual complications of influenza A infection in humans involve the skeletal muscles. Numerous cases of acute myopathy and/or rhabdomyolysis have been reported, particularly following the outbreak of pandemic influenza A(H1N1) in 2009. The pathogenesis of these influenza-associated myopathies (IAM) remains unkown, although the direct infection of muscle cells is suspected. Here, we studied the susceptibility of cultured human primary muscle cells to a 2009 pandemic and a 2008 seasonal influenza A(H1N1) isolate. Using cells from different donors, we found that differentiated muscle cells (i. e. myotubes) were highly susceptible to infection by both influenza A(H1N1) isolates, whereas undifferentiated cells (i. e. myoblasts) were partially resistant. The receptors for influenza viruses, alpha 2-6 and alpha 2-3 linked sialic acids, were detected on the surface of myotubes and myoblasts. Time line of viral nucleoprotein (NP) expression and nuclear export showed that the first steps of the viral replication cycle could take place in muscle cells. Infected myotubes and myoblasts exhibited budding virions and nuclear inclusions as observed by transmission electron microscopy and correlative light and electron microscopy. Myotubes, but not myoblasts, yielded infectious virus progeny that could further infect naive muscle cells after proteolytic treatment. Infection led to a cytopathic effect with the lysis of muscle cells, as characterized by the release of lactate dehydrogenase. The secretion of proinflammatory cytokines by muscle cells was not affected following infection. Our results are compatible with the hypothesis of a direct muscle infection causing rhabdomyolysis in IAM patients.
Journal Article
Evaluation of Tumor Budding in Primary Colorectal Cancer and Corresponding Liver Metastases Based on H&E and Pancytokeratin Staining
In colorectal cancer, tumor budding is associated with tumor progression and represents an additional prognostic factor in the TNM classification. Tumor buds can be found at the invasive front (peritumoral budding; PTB) and in the tumor center (intratumoral budding; ITB) of primary tumors. Previous studies have shown that tumor buds are also present in colorectal liver metastases (CRLM). Data on the prognostic and predictive role in this clinical context are still sparse and no standardized approach to evaluate budding in CRLM has been published so far. This study aimed to analyze and correlate perimetastatic (PMB) and intrametastatic budding (IMB) on H&E and pancytokeratin staining, compare it to budding results in corresponding primary tumors and to propose a standardized scoring system in CRLM as the basis for future studies. Tumor tissue of 81 primary tumors and 139 corresponding CRLM was used for ngTMA construction. For each primary tumor and metastasis, two punches from the center and two punches from the periphery from areas with highest tumor budding density were included. TMA slides were stained for H&E and pancytokeratin (Pan-CK). PTB, ITB, PMB, and IMB were analyzed and classified as bd1, bd2, and bd3 according to ITBCC guidelines. ITB and PTB as well as IMB and PMB showed significant correlation on H&E and Pan-CK staining. No correlation was found for tumor bud counts in primary tumors and corresponding metastases. The agreement for categorized tumor bud counts showed fair to good agreement for metastases and poor agreement for primary tumors between different classes on H&E and Pan-CK staining. Based on our results, tumor budding in primary tumors and CRLM seems to be different processes which might be the results of differing surrounding microenvironments. The evaluation of tumor budding in CRLM is challenging in cases without desmoplastic stroma reaction or intense perimetastatic ductular reaction. We therefore propose to evaluate tumor budding only in metastases with desmoplastic stroma reaction based on H&E staining since important morphological features are obscured on Pan-CK staining.
Journal Article
A method of genetic transformation and gene editing of succulents without tissue culture
Summary Succulents, valued for their drought tolerance and ornamental appeal, are important in the floriculture market. However, only a handful of succulent species can be genetically transformed, making it difficult to improve these plants through genetic engineering. In this study, we adapted the recently developed cut‐dip‐budding (CDB) gene delivery system to transform three previously recalcitrant succulent varieties – the dicotyledonous Kalanchoe blossfeldiana and Crassula arborescens and the monocotyledonous Sansevieria trifasciata. Capitalizing on the robust ability of cut leaves to regenerate shoots, these plants were successfully transformed by directly infecting cut leaf segments with the Agrobacterium rhizogenes strain K599. The transformation efficiencies were approximately 74%, 5% and 3.9%–7.8%, respectively, for K. blossfeldiana and C. arborescens and S. trifasciata. Using this modified CDB method to deliver the CRISPR/Cas9 construct, gene editing efficiency in K. blossfeldiana at the PDS locus was approximately 70%. Our findings suggest that succulents with shoot regeneration ability from cut leaves can be genetically transformed using the CDB method, thus opening up an avenue for genetic engineering of these plants.
Journal Article
Evaluation of Tumor Budding in Primary Colorectal Cancer and Corresponding Liver Metastases Based on H E and Pancytokeratin Staining
In colorectal cancer, tumor budding is associated with tumor progression and represents an additional prognostic factor in the TNM classification. Tumor buds can be found at the invasive front (peritumoral budding; PTB) and in the tumor center (intratumoral budding; ITB) of primary tumors. Previous studies have shown that tumor buds are also present in colorectal liver metastases (CRLM). Data on the prognostic and predictive role in this clinical context are still sparse and no standardized approach to evaluate budding in CRLM has been published so far. This study aimed to analyze and correlate perimetastatic (PMB) and intrametastatic budding (IMB) on H&E and pancytokeratin staining, compare it to budding results in corresponding primary tumors and to propose a standardized scoring system in CRLM as the basis for future studies. Tumor tissue of 81 primary tumors and 139 corresponding CRLM was used for ngTMA construction. For each primary tumor and metastasis, two punches from the center and two punches from the periphery from areas with highest tumor budding density were included. TMA slides were stained for H&E and pancytokeratin (Pan-CK). PTB, ITB, PMB, and IMB were analyzed and classified as bd1, bd2, and bd3 according to ITBCC guidelines. ITB and PTB as well as IMB and PMB showed significant correlation on H&E and Pan-CK staining. No correlation was found for tumor bud counts in primary tumors and corresponding metastases. The agreement for categorized tumor bud counts showed fair to good agreement for metastases and poor agreement for primary tumors between different classes on H&E and Pan-CK staining. Based on our results, tumor budding in primary tumors and CRLM seems to be different processes which might be the results of differing surrounding microenvironments. The evaluation of tumor budding in CRLM is challenging in cases without desmoplastic stroma reaction or intense perimetastatic ductular reaction. We therefore propose to evaluate tumor budding only in metastases with desmoplastic stroma reaction based on H&E staining since important morphological features are obscured on Pan-CK staining.
Journal Article
Influence of Time and Methods of Budding in ‘Khasi Mandarin’ (Citrus reticulata Blanco.) On Different Rootstocks in the Foothills of Arunachal Pradesh
North-East India is known for its high quality Khasi mandarin production. Multiplication of healthy and, true to the type of quality planting material is a necessity to strengthen the orchards and to boost its cultivation in the region. Different rootstocks viz., Rough lemon, Rangpur lime, Khasi mandarin, Volkamer lemon and Pummelo and different methods of budding (‘T’-budding and modified chip budding) were used for Khasi mandarin in February, March and April. Considering the method of budding, time and rootstocks the maximum bud take (93.33%), budding success (93.33%), sprout length (11.30 cm), sprout girth (2.14 mm), dry weight of sprout (2.95 g), number of leaves (7.97), Chlorophyll ‘a’ (0.34 mg/g), ‘b’ (0.50 mg/g) and total chlorophyll (0.86 mg/g) were recorded when modified chip budding was done on Rough lemon in February. Minimum days required to sprout (32.00) and maximum root diameter (8.23 mm) were observed on Khasi mandarin with modified chip budding done in February. Leaf perimeter (18.7 cm), root length (32.37 cm), fresh (22.13 g) and dry weight (16.22 g) of roots were maximum in Volkamer lemon with ‘T’-budding done during February. Thus, the study concluded that modified chip budding is the best method of budding to perform during February in Khasi mandarin using Rough lemon as rootstock with respect to bud take, budding success and growth of sprout.
Journal Article
Effect of Understumps Age on the Growth of Budding of Orange Plant (Citrus Sp.)
The research was conducted in the Garden of Development of Horticultural Seeds of West Java Province. The purpose of research to know the influence of age of rootstock and budding method to the success of citrus (Citrus sp). Research method using Factorial Randomized Block Design (RBD) with 2 factors. The first factor is the age of the Understump (rootstock) consisting of 5 levels, namely the stem age of 4, 6, 8, 10 and 12 months, while the second factor is the budding method consisting of 2 levels, namely T (T-budding) budding method and the budding (Chip-budding). The results showed that the treatment of budding method and the age of rootstock did not interact in all observation variables. The treatment of budding method and the age of rootstock independently had a very significant effect on the high percentage of shoots, no significant effect on the stem bud diameter and significantly affect the number of leaves. The best treatment on the method of budding is the method of Insert (Chip-Budding), while at the lowest rootstock is the best age of rootstock 6 months.
Journal Article
Nasopharyngeal carcinoma ecology theory: cancer as multidimensional spatiotemporal “unity of ecology and evolution” pathological ecosystem
Nasopharyngeal carcinoma (NPC) is a particular entity of head neck cancer that is generally regarded as a genetic disease with diverse intertumor and intratumor heterogeneity. This perspective review mainly outlines the up-to-date knowledge of cancer ecology and NPC progression, and presents a number of conceptual stepping-stones. At the beginning, I explicitly advocate that the nature of NPC (cancer) is not a genetic disease but an ecological disease: a multidimensional spatiotemporal \"unity of ecology and evolution\" pathological ecosystem. The hallmarks of cancer is proposed to act as ecological factors of population fitness. Subsequently, NPC cells are described as invasive species and its metastasis as a multidirectional ecological dispersal. The foundational ecological principles include intraspecific relationship (e.g. communication) and interspecific relationship (e.g. competition, predation, parasitism and mutualism) are interpreted to understand NPC progression. \"Mulberry-fish-ponds\" model can well illustrate the dynamic reciprocity of cancer ecosystem. Tumor-host interface is the ecological transition zone of cancer, and tumor buddings should be recognized as ecological islands separated from the mainland. It should be noted that tumor-host interface has a significantly molecular and functional edge effect because of its curvature and irregularity. Selection driving factors and ecological therapy including hyperthermia for NPC patients, and future perspectives in such field as \"ecological pathology\", \"multidimensional tumoriecology\" are also discussed. I advance that \"nothing in cancer evolution or ecology makes sense except in the light of the other\". The cancer ecology tree is constructed to comprehensively point out the future research direction. Taken together, the establishment of NPC ecology theory and cancer ecology tree might provide a novel conceptual framework and paradigm for our understanding of cancer complex causal process and potential preventive and therapeutic applications for patients.
Journal Article
Mechanics of Receptor-Mediated Endocytosis
Most viruses and bioparticles endocytosed by cells have characteristic sizes in the range of tens to hundreds of nanometers. The process of viruses entering and leaving animal cells is mediated by the binding interaction between ligand molecules on the viral capid and their receptor molecules on the cell membrane. How does the size of a bioparticle affect receptor-mediated endocytosis? Here, we study how a cell membrane containing diffusive mobile receptors wraps around a ligand-coated cylindrical or spherical particle. It is shown that particles in the size range of tens to hundreds of nanometers can enter or exit cells via wrapping even in the absence of clathrin or caveolin coats, and an optimal particles size exists for the smallest wrapping time. This model can also be extended to include the effect of clathrin coat. The results seem to show broad agreement with experimental observations.
Journal Article
Predictive power of tumour budding for lymph node metastasis in colorectal carcinomas: A retrospective study
Background & objectives: Tumour budding is a feature of epithelial-to-mesenchymal transformation that is characterized histologically within the tumour stroma by the presence of isolated cells or clusters of less than five cells which are different from the other malignant cells. This could be present around the invasive margin of the tumour, called peritumoural budding, or in the bulk of the tumour, called intratumoural budding. The aim of this study was to assess the predictive power of tumour budding for lymph node metastasis and its relationship with other features of tumour progression in colorectal carcinoma (CRC).
Methods: Preoperative colonoscopic biopsies and consecutive resection specimens from 80 patients of colorectal cancer were taken. In the biopsy, intratumoural budding was looked for and graded. In the resection, peritumoural budding was seen and graded along with other features such as grade of the tumour, lymphovascular emboli and tumour border configuration.
Results: Intratumoural budding was seen in 23 per cent (18/80) and peritumoural in 52 per cent (42/80) of cases. Intratumoural budding was associated with the presence of lymphovascular emboli (P=0.002) and irregular tumour border configuration (P=0.004). Peritumoural budding was also significantly associated with the presence of lymphovascular emboli and irregular margins (P < 0.001). Both intra- and peritumoural budding were not associated with the grade of the tumour. Both intra- and peritumoural budding had a significant association with lymph node metastasis (LNM) (P < 0.001).
Interpretation & conclusions: Our findings indicate that tumour budding in preoperative biopsy and resection specimens may predict a possibility of finding LNM in patients with CRC.
Journal Article