Restriction of Wolbachia bacteria in early embryogenesis of neotropical Drosophila species via ER-mediated autophagy

AbstractWolbachia bacteria are maternally transmitted intracellular microbes that are not only restricted to the reproductive organs but also found in various somatic tissues of their native hosts. The abundance of the endosymbiont in somatic tissues, usually a dead end for vertically transmitted bacteria, causes a multitude of effects on life history traits of their hosts, which are still not well understood. Thus, deciphering the host-symbiont interactions on a cellular level throughout a host’s lifecycle is of great importance to understand their homeostatic nature, persistence and spreading success. Using fluorescent and transmission electron microscopy, we conducted a comprehensive analysis of Wolbachia tropism in somatic and reproductive tissues of six Drosophila species at the intracellular level during host development. Our data uncovered diagnostic patterns of infections to embryonic primordial germ cells and to particular cells of somatic tissues in three different neotropical Drosophila species of the willistoni and saltans groups that have apparently evolved in both independently. We further found that restricted patterns of Wolbachia tropism are already determined in early fly embryogenesis. This is achieved via selective autophagy, and the restriction of infection is preserved through larval hatching and metamorphosis. We further uncovered tight interactions of Wolbachia with membranes of the endoplasmic reticulum, which might play a scaffolding role for autophagosome formation and subsequent elimination of the endosymbiont. Finally, by analyzing D. simulans lines transinfected with non-native Wolbachia, we uncovered that the host genetic background regulates tissue tropism of infection. Our data demonstrate a peculiar and novel mechanism to limit and spatially restrict bacterial infection in somatic tissues during a very early stage of host development.

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A Proof for the Possibility of Ce-Oxide from CMP Residuals in Si-Wafers by Analytical TEM

ISTFA 2012: Conference Proceedings from the 38th International Symposium for Testing and Failure Analysis ◽

10.31399/asm.cp.istfa2012p0347 ◽

2012 ◽

Author(s):

Wayne Zhao ◽

Liem Do Thanh ◽

Michael Gribelyuk ◽

Mary-Ann Zaitz ◽

Wing Lai

Keyword(s):

Technology Development ◽

Early Stage ◽

Chemical Mechanical Planarization ◽

Particle Cluster ◽

Limited Region ◽

Physical Evidence ◽

Analytical Tem ◽

Transmission Electron ◽

Oxide Particles

Abstract Inclusion of cerium (Ce) oxide particles as an abrasive into chemical mechanical planarization (CMP) slurries has become popular for wafer fabs below the 45nm technology node due to better polishing quality and improved CMP selectivity. Transmission electron microscopy (TEM) has difficulties finding and identifying Ce-oxide residuals due to the limited region of analysis unless dedicated efforts to search for them are employed. This article presents a case study that proved the concept in which physical evidence of Ce-rich particles was directly identified by analytical TEM during a CMP tool qualification in the early stage of 20nm node technology development. This justifies the need to setup in-fab monitoring for trace amounts of CMP residuals in Si-based wafer foundries. The fact that Cr resided right above the Ce-O particle cluster, further proved that the Ce-O particles were from the wafer and not introduced during the sample preparation.

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Ultrastructure of the female reproductive organs (ovary, vitellaria, and Mehlis' gland) of Halipegus eccentricus (Trematoda: Derogenidae)

Canadian Journal of Zoology ◽

10.1139/z86-334 ◽

1986 ◽

Vol 64 (10) ◽

pp. 2203-2212 ◽

Cited By ~ 22

Author(s):

Jon M. Holy ◽

Darwin D. Wittrock

Keyword(s):

Cell Types ◽

Reproductive Organs ◽

Golgi Body ◽

Digenetic Trematode ◽

Secretory Cells ◽

Cortical Granules ◽

Golgi Bodies ◽

Transmission Electron ◽

Vitelline Cells ◽

Female Reproductive Organs

The female reproductive organs (ovary, vitellaria, and Mehlis' gland) of the digenetic trematode Halipegus eccentricus were studied by transmission electron microscopy. Oocytes entered diplotene while in the ovary and produced cortical granules and lipid bodies. Vitelline cells produced large amounts of eggshell protein but no yolk bodies. Two types of Mehlis' gland secretory cells were present, distinguishable by the morphology of their rough endoplasmic reticulum, Golgi bodies, and secretory bodies, and by the persistence of recognizable secretory material within the ootype lumen after exocytosis. In an attempt to standardize the nomenclature regarding the cell types of the Mehlis' gland, a classification that takes into account these four criteria is proposed. Two basic types of Golgi body organization were noted for the cells of the female reproductive system: a stack of flattened cisternae (Mehlis' gland alpha cells) and spherical Golgi bodies with vesicular cisternae (oocytes, vitelline cells, and Mehlis' gland beta cells).

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Subcellular Investigation of Photosynthesis-Driven Carbon Assimilation in the Symbiotic Reef Coral Pocillopora damicornis

mBio ◽

10.1128/mbio.02299-14 ◽

2015 ◽

Vol 6 (1) ◽

Cited By ~ 57

Author(s):

Christophe Kopp ◽

Isabelle Domart-Coulon ◽

Stephane Escrig ◽

Bruno M. Humbel ◽

Michel Hignette ◽

...

Keyword(s):

Lipid Droplets ◽

Secondary Ion Mass Spectrometry ◽

Reef Coral ◽

Carbon Assimilation ◽

Isotopic Labeling ◽

Cellular Level ◽

Coral Tissue ◽

Pocillopora Damicornis ◽

Carbon And Nitrogen ◽

Transmission Electron

ABSTRACT Reef-building corals form essential, mutualistic endosymbiotic associations with photosynthetic Symbiodinium dinoflagellates, providing their animal host partner with photosynthetically derived nutrients that allow the coral to thrive in oligotrophic waters. However, little is known about the dynamics of these nutritional interactions at the (sub)cellular level. Here, we visualize with submicrometer spatial resolution the carbon and nitrogen fluxes in the intact coral-dinoflagellate association from the reef coral Pocillopora damicornis by combining nanoscale secondary ion mass spectrometry (NanoSIMS) and transmission electron microscopy with pulse-chase isotopic labeling using [13C]bicarbonate and [15N]nitrate. This allows us to observe that (i) through light-driven photosynthesis, dinoflagellates rapidly assimilate inorganic bicarbonate and nitrate, temporarily storing carbon within lipid droplets and starch granules for remobilization in nighttime, along with carbon and nitrogen incorporation into other subcellular compartments for dinoflagellate growth and maintenance, (ii) carbon-containing photosynthates are translocated to all four coral tissue layers, where they accumulate after only 15 min in coral lipid droplets from the oral gastroderm and within 6 h in glycogen granules from the oral epiderm, and (iii) the translocation of nitrogen-containing photosynthates is delayed by 3 h. IMPORTANCE Our results provide detailed in situ subcellular visualization of the fate of photosynthesis-derived carbon and nitrogen in the coral-dinoflagellate endosymbiosis. We directly demonstrate that lipid droplets and glycogen granules in the coral tissue are sinks for translocated carbon photosynthates by dinoflagellates and confirm their key role in the trophic interactions within the coral-dinoflagellate association.

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Light and electron microscopic investigations in the Dictyonemataceae (Basidiolichens)

Canadian Journal of Botany ◽

10.1139/b77-294 ◽

1977 ◽

Vol 55 (20) ◽

pp. 2565-2573 ◽

Cited By ~ 10

Author(s):

Robert D. Slocum ◽

Gary L. Floyd

Keyword(s):

Electron Microscopy ◽

Transmission Electron Microscopy ◽

Cellular Level ◽

Fungal Symbiont ◽

Electron Microscopic ◽

Basidiomycetous Fungus ◽

Transmission Electron ◽

Fungal Symbiosis ◽

Algal Host ◽

Microscopic Investigations

The nature of the association between the basidiomycetous mycobiont and the blue-green phycobiont in two species of the tropical basidiolichen Dictyonema was investigated using Nomarski light optics and scanning and transmission electron microscopy. Although members of this family may exhibit either a homoiomerous or heteromerous type of thallus organization, the fungus–alga relationship at the cellular level is remarkably consistent. Scytonema filaments are intimately associated with appressorial hyphae of the mycobiont and with extensive intracellular hyphae, which appear to be unrelated to the basidiomycetous fungal symbiont. This is the first report of a lichen displaying an apparent dual fungal symbiosis with the algal host. Association with the intracellular fungus produces no discernible damage to the phycobiont and apparently does not interfere with the symbiosis involving the basidiomycetous fungus.

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The fate and function of glycosphingolipid glucosylceramide

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2003.1266 ◽

2003 ◽

Vol 358 (1433) ◽

pp. 869-873 ◽

Cited By ~ 39

Author(s):

Gerrit van Meer ◽

Jasja Wolthoorn ◽

Sophie Degroote

Keyword(s):

Cellular Differentiation ◽

Membrane Lipids ◽

Early Stage ◽

Cellular Level ◽

Mature Stage ◽

Storage Diseases ◽

Head Groups ◽

Biological Studies ◽

Intracellular Membrane Transport ◽

And Function

In higher eukaryotes, glucosylceramide is the simplest member and precursor of a fascinating class of membrane lipids, the glycosphingolipids. These lipids display an astounding variation in their carbohydrate head groups, suggesting that glycosphingolipids serve specialized functions in recognition processes. It is now realized that they are organized in signalling domains on the cell surface. They are of vital importance as, in their absence, embryonal development is inhibited at an early stage. Remarkably, individual cells can live without glycolipids, perhaps because their survival does not depend on glycosphingolipid–mediated signalling mechanisms. Still, these cells suffer from defects in intracellular membrane transport. Various membrane proteins do not reach their intracellular destination, and, indeed, some intracellular organelles do not properly differentiate to their mature stage. The fact that glycosphingolipids are required for cellular differentiation suggests that there are human diseases resulting from defects in glycosphingolipid synthesis. In addition, the same cellular differentiation processes may be affected by defects in the degradation of glycosphingolipids. At the cellular level, the pathology of glycosphingolipid storage diseases is not completely understood. Cell biological studies on the intracellular fate and function of glycosphingolipids may open new ways to understand and defeat not only lipid storage diseases, but perhaps other diseases that have not been connected to glycosphingolipids so far.

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On the role of germ cells in planarian regeneration

Development ◽

10.1242/dev.55.1.53 ◽

1980 ◽

Vol 55 (1) ◽

pp. 53-63

Author(s):

V. Gremigni ◽

C. Miceli ◽

I. Puccinelli

Keyword(s):

Chromosome Number ◽

Germ Cells ◽

Primordial Germ Cells ◽

Karyological Analysis ◽

Blastema Formation ◽

Planarian Regeneration ◽

Somatic Tissues ◽

Caudal Area ◽

Complete Regeneration

Specimens from a polyploid biotype of Dugesia lugubris s.l. were used to clarify the role and fate of germ cells during planarian regeneration. These specimens provide a useful karyological marker because embryonic and somatic cells (3n = 12) can be easily distinguished from male (2n = 8) and female (6n = 24) germ cells by their chromosome number. We succeed in demonstrating how primordial germ cells participate in blastema formation and take part in rebuilding somatic tissues. This evidence was obtained by cutting each planarian specimen twice at appropriate levels. The first aimed to induce primordial germ cells to migrate to the wound. The second cut was performed after complete regeneration and aimed to obtain a blastema from a cephalic or caudal area devoid of gonads. A karyological analysis of mitotic cells present in each blastema obtained after the second cut provided evidence that cells, originally belonging to the germ lines, are still present in somatic tissues even months after complete regeneration. The role of primordial germ cells in planarian regeneration was finally discussed in relation to the phenomenon of metaplasia or transdifferentiation.

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Exogenous HGF Prevents Cardiomyocytes from Apoptosis after Hypoxia via Up-Regulating Cell Autophagy

Cellular Physiology and Biochemistry ◽

10.1159/000445592 ◽

2016 ◽

Vol 38 (6) ◽

pp. 2401-2413 ◽

Cited By ~ 19

Author(s):

Yunle Wang ◽

Jiabao Liu ◽

Zhiwen Tao ◽

Peng Wu ◽

Weili Cheng ◽

...

Keyword(s):

Therapeutic Strategy ◽

Protective Factor ◽

Ventricular Myocytes ◽

Early Stage ◽

Neonatal Rat ◽

Transmission Electron ◽

Neonatal Rat Heart ◽

Hoechst Staining ◽

Hepatocyte Growth

Background: Hepatocyte growth factor (HGF) is widely known as a protective factor in ischemic myocardium, however HGF sensitive cellular mechanism remained ill-defined. Autophagy at early stage of hypoxia has been demonstrated to play a role in protecting myocardium both in vivo and vitro. We performed this study to investigate the association between the protective effect of HGF and autophagy. Methods: Ventricular myocytes were isolated from neonatal rat heart (NRVMs). We evaluated cardiomyocytes apoptosis by Hoechst staining and flow cytometry. Autophagy was assessed by transmission electron microscope and mRFP-GFP-LC3 adenovirus infection. Mitochondrial membrane potential was estimated by JC-1 staining. Western blotting and ELISA assay were used to quantify protein concentrations. Results: We found that autophagy in NRVMs increased at early stage after hypoxia and HGF release was consistent with the change of autophagy. Exogenous HGF enhanced autophagy and decreased apoptosis, while neutralizing HGF yielded opposite effects. Besides, inhibition of autophagy increased apoptosis of myocytes. Furthermore, exogenous HGF induced Parkin, the marker of mitochondrial autophagy, indicating increased clearance of injured mitochondria. Conclusions: Our results revealed a potential mechanism in which exogenous HGF prevented NRVMs from apoptosis after hypoxia. Upregulation of Parkin through administration of exogenous HGF may be a potential therapeutic strategy ptotecting myocytes during ischemia.

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Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells

10.1101/866897 ◽

2019 ◽

Cited By ~ 1

Author(s):

Shivanand Hegde ◽

Denis Voronin ◽

Aitor Casas-Sanchez ◽

Miguel A. Saldaña ◽

Eva Heinz ◽

...

Keyword(s):

Zika Virus ◽

Control Strategies ◽

Fluorescent Microscopy ◽

Cellular Level ◽

Intracellular Bacteria ◽

Bacterial Symbionts ◽

Mosquito Cells ◽

Transmission Electron ◽

Bacterial Replication

AbstractMicrobiota within mosquitoes influence nutrition, immunity, fecundity, and the capacity to transmit pathogens. Despite their importance, we have a limited understanding of host-microbiota interactions, especially at the cellular level. It is evident bacterial symbionts that are localized within the midgut also infect other organs within the mosquito; however, the route these symbionts take to colonize other tissues is unknown. Here, utilizing the gentamicin protection assay, we showed that the bacterial symbionts Cedecea and Serratia have the capacity to invade and reside intracellularly within mosquito cells. Symbiotic bacteria were found within a vacuole and bacterial replication was observed in mosquito cell by transmission electron microscopy, indicating bacteria were adapted to the intracellular milieu. Using gene silencing, we determined that bacteria exploited host factors, including actin and integrin receptors, to actively invade mosquito cells. As microbiota can affect pathogens within mosquitoes, we examined the influence of intracellular symbionts on Zika virus (ZIKV) infection. Mosquito cells harbouring intracellular bacteria had significantly less ZIKV compared to uninfected cells or cells exposed to non-invasive bacteria. Intracellular bacteria were observed to substantially upregulate the Toll and IMD innate immune pathways, providing a possible mechanism mediating these anti-viral effects. Examining mono-axenically infected mosquitoes using transmission electron and fluorescent microscopy revealed that bacteria occupied an intracellular niche in vivo. Our results provided evidence that bacteria that associate with the midgut of mosquitoes have intracellular lifestyles which likely have implications for mosquito biology and pathogen infection. This study expands our understanding of host-microbiota interactions in mosquitoes, which is important as symbiont microbes are being exploited for vector control strategies.

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Perubahan nilai laktat serum dan nilai leukosit pasca penanganan pasien multitrauma

JURNAL BIOMEDIK (JBM) ◽

10.35790/jbm.8.2.2016.12703 ◽

2016 ◽

Vol 8 (2) ◽

Author(s):

Jimmy Koan ◽

Laurens T. B. Kalesaran ◽

Heber B. Sapan

Keyword(s):

Blood Lactate ◽

Analytical Study ◽

Early Stage ◽

Cellular Level ◽

Metabolic Changes ◽

T Test ◽

Laboratory Markers ◽

The Mean ◽

Paired T Test

Abstract: Available laboratory markers in the early stage of injury are very helpful for the clinicians to predict the diturbances in cellular level concerning prevention of early decompensation, therefore, vital condition of the patient can improve faster. Lactate and leucocyte levels are assumed as sensitive markers of metabolic changes that occur at the time of injury. This study aimed to obtain the changes of lactate and leukocyte levels in multitraumatic patients after resusitation at Prof. Dr. R. D. Kandou Hospital Manado from August to September 2015. This was an observational analytical study. The results showed that there were 36 multitraumatic patients in this study, consisted of 27 males and 9 females. One patient died during this study. The mean decrease of blood lactate was 1.4611 mmol/L, meanwhile, of leukocytes was 5582.2000/mm3. The paired T test showed very significant changes of blood lactate and leukocyte levels (P < 0.001) after resusitation. Conclusion: Achievement of resusitation and improvement in cellular level could be monitored by using lactate and leukocyte levels after resusitation of multitraumatic patients although the definitive aim of the trauma was not final yet.Kata kunci: lactate, leukocyte, multitraumatic patientsAbstrak: Tersedianya penanda laboratorik pada fase awal cedera dapat memudahkan klinisi memrediksi kelainan yang terjadi di tingkat sel untuk mencegah terjadinya fase dekompensasi secara dini sehingga dapat memperbaiki kondisi vital pasien dengan segera. Kadar laktat dan jumlah leukosit telah lama dianggap sebagai salah satu penanda yang sensitif terhadap perubahan metabolisme yang terjadi saat cedera. Penelitian ini bertujuan untuk mengetahui perubahan nilai serum laktat dan leukosit darah yang terjadi pada pasien multitrauma setelah penanganan di RSUP Prof. Dr. R. D. Kandou Manado sejak bulan Agustus sampai dengan September 2015. Jenis penelitian ini observasional analitik dengan desain potong lintang. Hasil penelitian memperlihatkan terdapat 36 pasien multitrauma, terdiri dari 27 laki-laki dan 9 perempuan. Selama penelitian didapatkan 1 pasien meninggal saat penanganan. Pada pasca penanganan, rerata penurunan nilai asam laktat darah sebesar 1,4611 mmol/L dan nilai leukosit sebesar 5582,2000/mm3. Hasil uji T berpasangan memperlihatkan perubahan nilai laktat darah dan leukosit pasca penanganan yang sangat bermakna (P < 0,001). Simpulan: Tercapainya resusitasi dan perbaikan di tingkat sel dapat dimonitor dari nilai laktat dan leukosit darah pasca penanganan pasien multitrauma walaupun penanganan belum sampai pada tujuan definitif trauma.Kata kunci: asam laktat, leukosit, multitrauma

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TArget-Responsive Subcellular Catabolism Analysis for Early-stage Antibody-Drug Conjugates Screening and Assessment

10.26434/chemrxiv.13309454 ◽

2020 ◽

Author(s):

Hua Sang ◽

Jiali Liu ◽

Fang Zhou ◽

Xiaofang Zhang ◽

Jingwei Zhang ◽

...

Keyword(s):

Quality Assessment ◽

Therapeutic Efficacy ◽

High Throughput Screening ◽

Early Stage ◽

Cellular Level ◽

Drug Conjugates ◽

Therapeutic Outcomes ◽

Lysosomal Proteolysis

Key events including antibody-antigen affinity, ADC internalization, trafficking and lysosomal proteolysis-mediated payload release combinatorially determine the therapeutic efficacy and safety for ADCs. Nevertheless, a universal technology that efficiently and conveniently evaluates the involvement of these above elements to ADC payload release and hence the final therapeutic outcomes for mechanistic studies and quality assessment is lacking. Considering the plethora of ADC candidates under development owing to the ever-evolving linker and drug chemistry, we developed a TArget-Responsive Subcellular Catabolism (TARSC) approach that measures catabolites kinetics for given ADCs and elaborates how each individual step ranging from antigen binding to lysosomal proteolysis affects ADC catabolism by targeted interferences. Using a commercial and a biosimilar ado-trastuzumab emtansine (T-DM1) as model ADCs, we recorded unequivocal catabolites kinetics for the two T-DM1s in the presence and absence of the targeted interferences. Their negligible differences in TARSC profiles fitting with their undifferentiated therapeutic outcomes suggested by in vitro viability assays and in vivo tumor growth assays, highlighting TARSC analysis as a good indicator of ADC efficacy and bioequivalency. Lastly, we demonstrated the use of TARSC in assessing payload release efficiency for a new Trastuzumab-toxin conjugate. Collectively, we demonstrated the use of TARSC in characterizing ADC catabolism at (sub)cellular level, and in systematically depicting whether given target proteins affect ADC payload release and hence therapeutic efficacy. We anticipate its future use in high-throughput screening, quality assessment and mechanistic understanding of ADCs for drug R&D before proceeding to costly in vivo experiments.

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