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2. Cytotoxic effects of ZnO nanoparticles on mouse testicular cells

Author

Han Z, Yan Q, Ge W, Liu ZG, Gurunathan S, De Felici M, Shen W, and Zhang XF

Subjects
ZnO nanoparticle, Sertoli cells, Leydig cells, mice, Medicine (General), R5-920
Abstract

Zhe Han,1,* Qi Yan,1,* Wei Ge,2 Zhi-Guo Liu,1 Sangiliyandi Gurunathan,3 Massimo De Felici,4 Wei Shen,2 Xi-Feng Zhang1 1College of Biological and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan, People’s Republic of China; 2Key Laboratory of Animal Reproduction and Germplasm Enhancement in Universities of Shandong, College of Animal Science and Technology, Qingdao Agricultural University, Qingdao, People’s Republic of China; 3Department of Stem Cell and Regenerative Biology, Konkuk University, Seoul, Republic of Korea; 4Department of Biomedicine and Prevention, University of Rome “Tor Vergata”, Rome, Italy *These authors contributed equally to this work Background: Nanoscience and nanotechnology are developing rapidly, and the applications of nanoparticles (NPs) have been found in several fields. At present, NPs are widely used in traditional consumer and industrial products, however, the properties and safety of NPs are still unclear and there are concerns about their potential environmental and health effects. The aim of the present study was to investigate the potential toxicity of ZnO NPs on testicular cells using both in vitro and in vivo systems in a mouse experimental model. Methods: ZnO NPs with a crystalline size of 70 nm were characterized with various analytical techniques, including ultraviolet-visible spectroscopy, Fourier transform infrared spectroscopy, X-ray diffraction, transmission electron microscopy, and atomic force microscopy. The cytotoxicity of the ZnO NPs was examined in vitro on Leydig cell and Sertoli cell lines, and in vivo on the testes of CD1 mice injected with single doses of ZnO NPs.Results: ZnO NPs were internalized by Leydig cells and Sertoli cells, and this resulted in cytotoxicity in a time- and dose-dependent manner through the induction of apoptosis. Apoptosis likely occurred as a consequence of DNA damage (detected as γ-H2AX and RAD51 foci) caused by increase in reactive oxygen species associated with loss of mitochondrial membrane potential. In addition, injection of ZnO NPs in male mice caused structural alterations in the seminiferous epithelium and sperm abnormalities.Conclusion: These results demonstrate that ZnO NPs have the potential to induce apoptosis in testicular cells likely through DNA damage caused by reactive oxygen species, with possible adverse consequences for spermatogenesis and therefore, male fertility. This suggests that evaluating the potential impacts of engineered NPs is essential prior to their mass production, to address both the environmental and human health concerns and also to develop sustainable and safer nanomaterials. Keywords: ZnO nanoparticle, Sertoli cells, Leydig cells, mice

Published
2016

5. Analysis of Secreted Proteins from Prepubertal Ovarian Tissues Exposed In Vitro to Cisplatin and LH

Author

Marcozzi, S., Ciccosanti, F., Fimia, G. M., Piacentini, M., Caggiano, C., Sette, C., De Felici, M., Klinger, F. G., Caggiano C., Sette C. (ORCID:0000-0003-2864-8266), Marcozzi, S., Ciccosanti, F., Fimia, G. M., Piacentini, M., Caggiano, C., Sette, C., De Felici, M., Klinger, F. G., Caggiano C., and Sette C. (ORCID:0000-0003-2864-8266)

Abstract

It is well known that secreted and exosomal proteins are associated with a broad range of physiological processes involving tissue homeostasis and differentiation. In the present paper, our purpose was to characterize the proteome of the culture medium in which the oocytes within the primordial/primary follicles underwent apoptosis induced by cisplatin (CIS) or were, for the most part, protected by LH against the drug. To this aim, prepubertal ovarian tissues were cultured under control and in the presence of CIS, LH, and CIS + LH. The culture media were harvested after 2, 12, and 24 h from chemotherapeutic drug treatment and analyzed by liquid chromatography–mass spectrometry (LC-MS). We found that apoptotic conditions generated by CIS in the cultured ovarian tissues and/or oocytes are reflected in distinct changes in the extracellular microenvironment in which they were cultured. These changes became evident mainly from 12 h onwards and were characterized by the inhibition or decreased release of a variety of compounds, such as the proteases Htra1 and Prss23, the antioxidants Prdx2 and Hbat1, the metabolic regulators Ldha and Pkm, and regulators of apoptotic pathways such as Tmsb4x. Altogether, these results confirm the biological relevance of the LH action on prepuberal ovaries and provide novel information about the proteins released by the ovarian tissues exposed to CIS and LH in the surrounding microenvironment. These data might represent a valuable resource for future studies aimed to clarify the effects and identify biomarkers of these compounds’ action on the developing ovary.

Published
2022

10. Cell Death in Mammalian Ovary

Author

Vázquez-Nin, Gerardo H., primary, Escobar, María Luisa, additional, De Felici, M., additional, Echeverría, Olga Margarita, additional, and Klinger, Francesca Gioia, additional

Published
2011
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16. Purification of mouse primordial germ cells by miniMACS magnetic separation system

Author

Pesce, M. and De Felici, M.

Subjects
Mice -- Anatomy, Embryology -- Research, Magnetic separators -- Usage, Germ cells -- Research, Cell separation -- Methods, Biological sciences
Abstract

The magnetic cell sorter miniMACS is used to isolate rare primordial germ cells (PGCs) from mouse embryos between 10.5 and 13.5 days post coitum. The labeled germ cells retained in the magnetic column are easily eluted and then can be used immediately for biochemical studies or cultured in appropriate in vitro systems for future germ cell experiments.

Published
1995

18. The cyto-protective effects of LH on ovarian reserve and female fertility during exposure to gonadotoxic alkylating agents in an adult mouse model.

Author

Castillo, L M Del, Buigues, A, Rossi, V, Soriano, M J, Martinez, J, Felici, M De, Lamsira, H K, Rella, F Di, Klinger, F G, Pellicer, A, Herraiz, S, Del Castillo, L M, De Felici, M, and Di Rella, F

Subjects
OVARIAN reserve, OVARIES, LABORATORY mice, ALKYLATING agents, INDUCED ovulation, FERTILITY preservation, ADULTS, WARNING labels, RESEARCH, ANIMAL experimentation, RESEARCH methodology, MEDICAL cooperation, EVALUATION research, COMPARATIVE studies, RESEARCH funding, MICE
Abstract

Study Question: Does LH protect mouse oocytes and female fertility from alkylating chemotherapy?Summary Answer: LH treatment before and during chemotherapy prevents detrimental effects on follicles and reproductive lifespan.What Is Known Already: Chemotherapies can damage the ovary, resulting in premature ovarian failure and reduced fertility in cancer survivors. LH was recently suggested to protect prepubertal mouse follicles from chemotoxic effects of cisplatin treatment.Study Design, Size, Duration: This experimental study investigated LH effects on primordial follicles exposed to chemotherapy. Seven-week-old CD-1 female mice were randomly allocated to four experimental groups: Control (n = 13), chemotherapy (ChT, n = 15), ChT+LH-1x (n = 15), and ChT+LH-5x (n = 8). To induce primary ovarian insufficiency (POI), animals in the ChT and ChT+LH groups were intraperitoneally injected with 120 mg/kg of cyclophosphamide and 12 mg/kg of busulfan, while control mice received vehicle. For LH treatment, the ChT+LH-1x and ChT+LH-5x animals received a 1 or 5 IU LH dose, respectively, before chemotherapy, then a second LH injection administered with chemotherapy 24 h later. Then, two animals/group were euthanized at 12 and 24 h to investigate the early ovarian response to LH, while remaining mice were housed for 30 days to evaluate short- and long-term reproductive outcomes. The effects of LH and chemotherapy on growing-stage follicles were analyzed in a parallel experiment. Seven-week-old NOD-SCID female mice were allocated to control (n = 5), ChT (n = 5), and ChT+LH-1x (n = 6) groups. Animals were treated as described above, but maintained for 7 days before reproductive assessment.Participants/materials, Setting, Methods: In the first experiment, follicular damage (phosphorylated H2AX histone (γH2AX) staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay), apoptotic biomarkers (western blot), and DNA repair pathways (western blot and RT-qPCR) were assessed in ovaries collected at 12 and 24 h to determine early ovarian responses to LH. Thirty days after treatments, remaining mice were stimulated (10 IU of pregnant mare serum gonadotropin (PMSG) and 10 IU of hCG) and mated to collect ovaries, oocytes, and embryos. Histological analysis was performed on ovarian samples to investigate follicular populations and stromal status, and meiotic spindle and chromosome alignment was measured in oocytes by confocal microscopy. Long-term effects were monitored by assessing pregnancy rate and litter size during six consecutive breeding attempts. In the second experiment, mice were stimulated and mated 7 days after treatments and ovaries, oocytes, and embryos were collected. Follicular numbers, follicular protection (DNA damage and apoptosis by H2AX staining and TUNEL assay, respectively), and ovarian stroma were assessed. Oocyte quality was determined by confocal analysis.Main Results and the Role Of Chance: LH treatment was sufficient to preserve ovarian reserve and follicular development, avoid atresia, and restore ovulation and meiotic spindle configuration in mature oocytes exposed at the primordial stage. LH improved the cumulative pregnancy rate and litter size in six consecutive breeding rounds, confirming the potential of LH treatment to preserve fertility. This protective effect appeared to be mediated by an enhanced early DNA repair response, via homologous recombination, and generation of anti-apoptotic signals in the ovary a few hours after injury with chemotherapy. This response ameliorated the chemotherapy-induced increase in DNA-damaged oocytes and apoptotic granulosa cells. LH treatment also protected growing follicles from chemotherapy. LH reversed the chemotherapy-induced depletion of primordial and primary follicular subpopulations, reduced oocyte DNA damage and granulosa cell apoptosis, restored mature oocyte cohort size, and improved meiotic spindle properties.Large Scale Data: N/A.Limitations, Reasons For Caution: This was a preliminary study performed with mouse ovarian samples. Therefore, preclinical research with human samples is required for validation.Wider Implications Of the Findings: The current study tested if LH could protect the adult mouse ovarian reserve and reproductive lifespan from alkylating chemotherapy. These findings highlight the therapeutic potential of LH as a complementary non-surgical strategy for preserving fertility in female cancer patients.Study Funding/competing Interest(s): This study was supported by grants from the Regional Valencian Ministry of Education (PROMETEO/2018/137), the Spanish Ministry of Science and Innovation (CP19/00141), and the Spanish Ministry of Education, Culture and Sports (FPU16/05264). The authors declare no conflict of interest. [ABSTRACT FROM AUTHOR]

Published
2021
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19. Elementi di istologia e cenni di embriologia

Author

Adamo, S., Bernardini, N., Boitani, C., Bonsi, L., Bouché, M., Brun, P., Canipari, R., Castriconi, R., Castrogiovanni, P., Ciccarelli, C., DE ANGELIS, L., D’Alessio, A., DE CESARIS, P., DE FELICI, M., DE MATTEI, M., Desiderio, V., DI ROSA, M., Dolfi, A., Dupont, S., Fazi, F., Filippini, A., Gagliano, N., Grano, M., Imbesi, R., Marcenaro, E., Musarò, A., Nervi, C., Papaccio, G. P., Ricci, G., Riccioli, A., Salustri, A., Scicchitano, B. M., Sivori, S., Tirino, V., Vicini, E., Virgintino, D., and Ziparo, E.

Published
2019

23. ISTOLOGIA di Monesi 7

Author

Adamo, S., De Felici, M., Dolfi, A., Filippini, A., Grano, M., Musarò, A., Nervi, C., Papaccio, G., Salustri, A., Ziparo, E., Bernardini, N., Boitani, C., Bonsi, L., Bouchè, M., Brun, P., Canipari, R., Castriconi, R., Castrogiovanni, P., Ciccarelli, C., De Cesaris, P., De Mattei, M., Desiderio, V., Dupont, S., Fazi, F., Gagliano, N., Imbesi, R., Marcenaro, E., Riccioli, A., Sivori, S., Tirino, V., Vicini, E., and Virgintino, D.

Subjects
Citologia Istologia
Published
2018

25. Il Citoscheletro

Author

Adamo, S., DE FELICI, M., Dolfi, A., Filippini, A., Grano, M., Musarò, A., Nervi, C., Papaccio, G., Salustri, A., Ziparo, E., Bernardini, N., Boitani, C., Bonsi, L., Bouché, M., Brun, P., Canipari, R., Castriconi, R., Castrogiovanni, P., Ciccarelli, C., DE CESARIS, P., DE MATTEI, M., Desiderio, V., Dupont, S., Fazi, F., Gagliano, N., Imbesi, R., Marcenaro, E., Riccioli, A., Sivori, S., Tirino, V., Vicini, E., and Virgintino, D.

Published
2018

26. Melatonin protects prepuberal testis from deleterious effects of bisphenol A or diethylhexyl phthalate by preserving H3K9 methylation

Author

Zhang, T., Zhou, Y., Li, L., Zhao, Y., De Felici, M., Reiter, R.J., Shen, W., Zhang, T., Zhou, Y., Li, L., Zhao, Y., De Felici, M., Reiter, R.J., and Shen, W.

Abstract

A growing number of couples experience fertility issues with almost half being due to malefactors. The exposure to toxic environmental contaminants, such as endocrine disruptors (EDs), has been shown to negatively affect male fertility. EDs are present in the environment, and exposure to these toxins results in the failure of spermatogenesis. The deleterious effects of EDs on spermatogenesis have been well documented, whereas improvement of infertility associated with spermatogenesis defects remains a great challenge. Herein, we report that in vitro exposure of prepuberal mouse testes to two well-known endocrine disruptors (EDs), bisphenol A (BPA) or diethylhexyl phthalate (DEHP), impairs spermatogenesis with perturbing self-renewal, spermatogonia activity, and meiosis. Evidence indicates that such effects are likely due, at least in part, to decreased G9a-dependent H3K9 di-methylation. Of note, we found that melatonin (MLT) protected the testis from the negative ED impacts with preserving spermatogonia stem and meiotic cells, along with maintaining normal H3K9 di-methylation in these cells. Taken together, this work documents that BPA and EDHP adversely affect prepuberal spermatogenesis and perturb crucial epigenetic activities in male germ cells and highlight the protective ability of MLT.

Published
2018

27. Monte Carlo code comparison of dose delivery prediction for Microbeam Radiation Therapy.

Author

de Felici, M, Siegbahn, E, Spiga, J, Hanson, A, Felici, R, Ferrero, C, Tartari, A, Gambaccini, M, Keyrilaeinen, J, Braeuer-Krisch, E, Randaccio, P, Bravin, A, BA Verhaegen, F, Seuntjens, J, de Felici M., Siegbahn E. A., Spiga J., Hanson A. L., Felici R., Ferrero C., Tartari A., Gambaccini M., Keyrilaeinen J., Braeuer-Krisch E., Randaccio P., Bravin A, BA Verhaegen F, Seuntjens J, de Felici, M, Siegbahn, E, Spiga, J, Hanson, A, Felici, R, Ferrero, C, Tartari, A, Gambaccini, M, Keyrilaeinen, J, Braeuer-Krisch, E, Randaccio, P, Bravin, A, BA Verhaegen, F, Seuntjens, J, de Felici M., Siegbahn E. A., Spiga J., Hanson A. L., Felici R., Ferrero C., Tartari A., Gambaccini M., Keyrilaeinen J., Braeuer-Krisch E., Randaccio P., Bravin A, BA Verhaegen F, and Seuntjens J

Abstract

Preclinical Microbeam Radiation Therapy (MRT) research programs are carried out at the European Synchrotron Radiation Facility (ESRF) and at a few other synchrotron facilities. MRT needs an accurate evaluation of the doses delivered to biological tissues for carrying out pre-clinical studies. This point is crucial for determining the effect induced by changing any of the physical irradiation parameters. The doses of interest in MRT are normally calculated using Monte Carlo (MC) methods. A few MC packages have been used in the last decade for MRT dose evaluations in independent studies. The aim of this investigation is to provide a preliminary basis to perform a systematic comparison of the dose results obtained, under identical irradiation conditions and for the same scoring geometries with the following five MC codes: EGS4, PENELOPE, GEANT4, EGSnrc, and MCNPX. Dose profiles have been calculated in an in-depth region of cylindrical phantoms made of water or PMMA. Beams in both cylindrical and planar geometry have been considered. This comparison shows an overall agreement among the different codes although minor differences occur, which need further investigations. © 2008 IOP Publishing Ltd.

Published
2008

28. Effects induced by the fine structure of biological tissues on microbeam dose deposition: A numerical study

Author

De Felici, M, Tartari, A, Taibi, A, Gambaccini, M, Felici, R, Bravin, A, Ferrero, C, Finet, S, De Felici M, Tartari A, Taibi A, Gambaccini M, Felici R, Bravin A, Ferrero C, Finet S, De Felici, M, Tartari, A, Taibi, A, Gambaccini, M, Felici, R, Bravin, A, Ferrero, C, Finet, S, De Felici M, Tartari A, Taibi A, Gambaccini M, Felici R, Bravin A, Ferrero C, and Finet S

Abstract

In this work small angle coherent scattering data are presented for some biological tissues. The effect of low angle diffusion on the micro-beams dose distributions are assessed by means of a dedicated version of the EGS4 code. The atomic form factor tabulations were upgraded in EGS4 to include experimental values of compounds instead of obtaining the linear differential scattering coefficient from a simple weighted sum of the elemental components. In this manner, one includes the effects induced by both the large-scale arrangement of the sample structure and the molecular interference. It is shown that the inclusion of the measured form factors and cross-sections in the photon transport calculations induce not negligible effects in the dose deposited around the microbeams. The fact of not taking into account small angle scattering cross-sections may lead to underestimating the number of scattered photons reaching zones not directly illuminated. At the center of a microbeam planar array the effect can be as much as 100% for X-ray photons of 50 keV.

Published
2005

29. Di-(2-ethylhexyl) phthalate and bisphenol A exposure impairs mouse primordial follicle assembly in vitro

Author

Zhang, T, Li, L, Qin, X, Zhou, Y, Zhang, X, Wang, L, DE FELICI, M, Chen, H, Qin, G, and Shen, W

Subjects
Cells, Messenger, Real-Time Polymerase Chain Reaction, SCID, Immunoenzyme Techniques, Mice, bisphenol-A, Oogenesis, Lhx8, Phenols, Ovarian Follicle, Plasticizers, Diethylhexyl Phthalate, Animals, Benzhydryl Compounds, Reverse Transcriptase Polymerase Chain Reaction, RNA, Messenger, Animals, Newborn, Blotting, Western, di-(2-ethylhexyl) phthalate, primordial follicle, Cells, Cultured, Free Radical Scavengers, DNA methylation, Oocytes, Mice, SCID, Female, Cultured, Settore BIO/17, Blotting, Newborn, RNA, Western
Published
2014

31. Guidelines for the use and interpretation of assays for monitoring autophagy

Author

Klionsky, D.J. Abdalla, F.C. Abeliovich, H. Abraham, R.T. Acevedo-Arozena, A. Adeli, K. Agholme, L. Agnello, M. Agostinis, P. Aguirre-Ghiso, J.A. Ahn, H.J. Ait-Mohamed, O. Ait-Si-Ali, S. Akematsu, T. Akira, S. Al-Younes, H.M. Al-Zeer, M.A. Albert, M.L. Albin, R.L. Alegre-Abarrategui, J. Aleo, M.F. Alirezaei, M. Almasan, A. Almonte-Becerril, M. Amano, A. Amaravadi, R. Amarnath, S. Amer, A.O. Andrieu-Abadie, N. Anantharam, V. Ann, D.K. Anoopkumar-Dukie, S. Aoki, H. Apostolova, N. Arancia, G. Aris, J.P. Asanuma, K. Asare, N.Y.O. Ashida, H. Askanas, V. Askew, D.S. Auberger, P. Baba, M. Backues, S.K. Baehrecke, E.H. Bahr, B.A. Bai, X.-Y. Bailly, Y. Baiocchi, R. Baldini, G. Balduini, W. Ballabio, A. Bamber, B.A. Bampton, E.T.W. Bánhegyi, G. Bartholomew, C.R. Bassham, D.C. Bast Jr., R.C. Batoko, H. Bay, B.-H. Beau, I. Béchet, D.M. Begley, T.J. Behl, C. Behrends, C. Bekri, S. Bellaire, B. Bendall, L.J. Benetti, L. Berliocchi, L. Bernardi, H. Bernassola, F. Besteiro, S. Bhatia-Kissova, I. Bi, X. Biard-Piechaczyk, M. Blum, J.S. Boise, L.H. Bonaldo, P. Boone, D.L. Bornhauser, B.C. Bortoluci, K.R. Bossis, I. Bost, F. Bourquin, J.-P. Boya, P. Boyer-Guittaut, M. Bozhkov, P.V. Brady, N.R. Brancolini, C. Brech, A. Brenman, J.E. Brennand, A. Bresnick, E.H. Brest, P. Bridges, D. Bristol, M.L. Brookes, P.S. Brown, E.J. Brumell, J.H. Brunetti-Pierri, N. Brunk, U.T. Bulman, D.E. Bultman, S.J. Bultynck, G. Burbulla, L.F. Bursch, W. Butchar, J.P. Buzgariu, W. Bydlowski, S.P. Cadwell, K. Cahová, M. Cai, D. Cai, J. Cai, Q. Calabretta, B. Calvo-Garrido, J. Camougrand, N. Campanella, M. Campos-Salinas, J. Candi, E. Cao, L. Caplan, A.B. Carding, S.R. Cardoso, S.M. Carew, J.S. Carlin, C.R. Carmignac, V. Carneiro, L.A.M. Carra, S. Caruso, R.A. Casari, G. Casas, C. Castino, R. Cebollero, E. Cecconi, F. Celli, J. Chaachouay, H. Chae, H.-J. Chai, C.-Y. Chan, D.C. Chan, E.Y. Chang, R.C.-C. Che, C.-M. Chen, C.-C. Chen, G.-C. Chen, G.-Q. Chen, M. Chen, Q. Chen, S.S.-L. Chen, W. Chen, X. Chen, X. Chen, X. Chen, Y.-G. Chen, Y. Chen, Y. Chen, Y.-J. Chen, Z. Cheng, A. Cheng, C.H.K. Cheng, Y. Cheong, H. Cheong, J.-H. Cherry, S. Chess-Williams, R. Cheung, Z.H. Chevet, E. Chiang, H.-L. Chiarelli, R. Chiba, T. Chin, L.-S. Chiou, S.-H. Chisari, F.V. Cho, C.H. Cho, D.-H. Choi, A.M.K. Choi, D. Choi, K.S. Choi, M.E. Chouaib, S. Choubey, D. Choubey, V. Chu, C.T. Chuang, T.-H. Chueh, S.-H. Chun, T. Chwae, Y.-J. Chye, M.-L. Ciarcia, R. Ciriolo, M.R. Clague, M.J. Clark, R.S.B. Clarke, P.G.H. Clarke, R. Codogno, P. Coller, H.A. Colombo, M.I. Comincini, S. Condello, M. Condorelli, F. Cookson, M.R. Coombs, G.H. Coppens, I. Corbalan, R. Cossart, P. Costelli, P. Costes, S. Coto-Montes, A. Couve, E. Coxon, F.P. Cregg, J.M. Crespo, J.L. Cronjé, M.J. Cuervo, A.M. Cullen, J.J. Czaja, M.J. D'Amelio, M. Darfeuille-Michaud, A. Davids, L.M. Davies, F.E. De Felici, M. De Groot, J.F. De Haan, C.A.M. De Martino, L. De Milito, A. De Tata, V. Debnath, J. Degterev, A. Dehay, B. Delbridge, L.M.D. Demarchi, F. Deng, Y.Z. Dengjel, J. Dent, P. Denton, D. Deretic, V. Desai, S.D. Devenish, R.J. Di Gioacchino, M. Di Paolo, G. Di Pietro, C. Díaz-Araya, G. Díaz-Laviada, I. Diaz-Meco, M.T. Diaz-Nido, J. Dikic, I. Dinesh-Kumar, S.P. Ding, W.-X. Distelhorst, C.W. Diwan, A. Djavaheri-Mergny, M. Dokudovskaya, S. Dong, Z. Dorsey, F.C. Dosenko, V. Dowling, J.J. Doxsey, S. Dreux, M. Drew, M.E. Duan, Q. Duchosal, M.A. Duff, K. Dugail, I. Durbeej, M. Duszenko, M. Edelstein, C.L. Edinger, A.L. Egea, G. Eichinger, L. Eissa, N.T. Ekmekcioglu, S. El-Deiry, W.S. Elazar, Z. Elgendy, M. Ellerby, L.M. Er Eng, K. Engelbrecht, A.-M. Engelender, S. Erenpreisa, J. Escalante, R. Esclatine, A. Eskelinen, E.-L. Espert, L. Espina, V. Fan, H. Fan, J. Fan, Q.-W. Fan, Z. Fang, S. Fang, Y. Fanto, M. Fanzani, A. Farkas, T. Farré, J.-C. Faure, M. Fechheimer, M. Feng, C.G. Feng, J. Feng, Q. Feng, Y. Fésüs, L. Feuer, R. Figueiredo-Pereira, M.E. Fimia, G.M. Fingar, D.C. Finkbeiner, S. Finkel, T. Finley, K.D. Fiorito, F. Fisher, E.A. Fisher, P.B. Flajolet, M. Florez-McClure, M.L. Florio, S. Fon, E.A. Fornai, F. Fortunato, F. Fotedar, R. Fowler, D.H. Fox, H.S. Franco, R. Frankel, L.B. Fransen, M. Fuentes, J.M. Fueyo, J. Fujii, J. Fujisaki, K. Fujita, E. Fukuda, M. Furukawa, R.H. Gaestel, M. Gailly, P. Gajewska, M. Galliot, B. Galy, V. Ganesh, S. Ganetzky, B. Ganley, I.G. Gao, F.-B. Gao, G.F. Gao, J. Garcia, L. Garcia-Manero, G. Garcia-Marcos, M. Garmyn, M. Gartel, A.L. Gatti, E. Gautel, M. Gawriluk, T.R. Gegg, M.E. Geng, J. Germain, M. Gestwicki, J.E. Gewirtz, D.A. Ghavami, S. Ghosh, P. Giammarioli, A.M. Giatromanolaki, A.N. Gibson, S.B. Gilkerson, R.W. Ginger, M.L. Ginsberg, H.N. Golab, J. Goligorsky, M.S. Golstein, P. Gomez-Manzano, C. Goncu, E. Gongora, C. Gonzalez, C.D. Gonzalez, R. González-Estévez, C. González-Polo, R.A. Gonzalez-Rey, E. Gorbunov, N.V. Gorski, S. Goruppi, S. Gottlieb, R.A. Gozuacik, D. Granato, G.E. Grant, G.D. Green, K.N. Gregorc, A. Gros, F. Grose, C. Grunt, T.W. Gual, P. Guan, J.-L. Guan, K.-L. Guichard, S.M. Gukovskaya, A.S. Gukovsky, I. Gunst, J. Gustafsson, A.B. Halayko, A.J. Hale, A.N. Halonen, S.K. Hamasaki, M. Han, F. Han, T. Hancock, M.K. Hansen, M. Harada, H. Harada, M. Hardt, S.E. Harper, J.W. Harris, A.L. Harris, J. Harris, S.D. Hashimoto, M. Haspel, J.A. Hayashi, S.-I. Hazelhurst, L.A. He, C. He, Y.-W. Hébert, M.-J. Heidenreich, K.A. Helfrich, M.H. Helgason, G.V. Henske, E.P. Herman, B. Herman, P.K. Hetz, C. Hilfiker, S. Hill, J.A. Hocking, L.J. Hofman, P. Hofmann, T.G. Höhfeld, J. Holyoake, T.L. Hong, M.-H. Hood, D.A. Hotamisligil, G.S. Houwerzijl, E.J. Høyer-Hansen, M. Hu, B. Hu, C.-A.A. Hu, H.-M. Hua, Y. Huang, C. Huang, J. Huang, S. Huang, W.-P. Huber, T.B. Huh, W.-K. Hung, T.-H. Hupp, T.R. Hur, G.M. Hurley, J.B. Hussain, S.N.A. Hussey, P.J. Hwang, J.J. Hwang, S. Ichihara, A. Ilkhanizadeh, S. Inoki, K. Into, T. Iovane, V. Iovanna, J.L. Ip, N.Y. Isaka, Y. Ishida, H. Isidoro, C. Isobe, K.-I. Iwasaki, A. Izquierdo, M. Izumi, Y. 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Abstract

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field. © 2012 Landes Bioscience.

Published
2012

42. EMBRIOLOGIA UMANA

Author

DE FELICI, M, Boitani, C, Bouchè, M, Canipari, R, Dolfi, A, Filippini, A, Musaro, A, Papaccio, G, and Salustri, A

Subjects
Settore BIO/17
Published
2009

48. Structural characterization of the human cerebral myelin sheath by small angle x-ray scattering

Author

De Felici, M, Felici, R, Ferrero, C, Tartari, A, Gambaccini, M, and Finet, S

Subjects
COHERENT SCATTERING, MEMBRANE-STRUCTURE, NERVE MYELIN, IN-VITRO, HUMAN BREAST-TISSUE
Abstract

Myelin is a multi-lamellar membrane surrounding neuronal axons and increasing their conduction velocity. When investigated by small-angle x-ray scattering (SAXS), the lamellar quasi-periodical arrangement of the myelin sheath gives rise to distinct peaks, which allow the determination of its molecular organization and the dimensions of its substructures. In this study we report on the myelin sheath structural determination carried out on a set of human brain tissue samples coming from surgical biopsies of two patients: a man around 60 and a woman nearly 90 years old. The samples were extracted either from white or grey cerebral matter and did not undergo any manipulation or chemical-physical treatment, which could possibly have altered their structure, except dipping them into a formalin solution for their conservation. Analysis of the scattered intensity from white matter of intact human cerebral tissue allowed the evaluation not only of the myelin sheath periodicity but also of its electronic charge density profile. In particular, the thicknesses of the cytoplasm and extracellular regions were established, as well as those of the hydrophilic polar heads and hydrophobic tails of the lipid bilayer. SAXS patterns were measured at several locations on each sample in order to establish the statistical variations of the structural parameters within a single sample and among different samples. This work demonstrates that a detailed structural analysis of the myelin sheath can also be carried out in randomly oriented samples of intact human white matter, which is of importance for studying the aetiology and evolution of the central nervous system pathologies inducing myelin degeneration.

Published
2008

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